Heart Rot and Root Rot in Tropical Acacia Plantations

Heart rot and root rot in tropical Acacia plantations

Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006

Editors: Karina Potter, Anto Rimbawanto and Chris Beadle

Australian Centre for International Agricultural Research Canberra 2006 The Australian Centre for International Agricultural Research (ACIAR) was established in June 1982 by an Act of the Australian Parliament. Its mandate is to help identify agricultural problems in developing countries and to commission collaborative research between Australian and developing country researchers in fields where Australia has a special research competence. Where trade names are used this constitutes neither endorsement of nor discrimination against any product by the Centre.

ACIAR PROCEEDINGS SERIES This series of publications includes the full proceedings of research workshops or symposia organised or supported by ACIAR. Numbers in this series are distributed internationally to selected individuals and scientific institutions.

Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124, 92p.

ISBN 1 86320 507 1 print ISBN 1 86320 510 1 online

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Foreword

Fast-growing hardwood plantations are increasingly important to the economies of many countries around the Pacific rim, including Australia, Indonesia and the Philippines. While the initial emphasis in most countries has been on the market for fibre, pulp and paper, there is now increasing interest in using the species for the production of solid-wood products. Indonesia has been at the forefront of the domestication of Australian acacias, especially Acacia mangium, and there is now an impressive estate of over one million hectares of acacias in the country. This resource is progressively replacing natural forest as a source of fibre for Indonesia’s large paper-producing industry. Acacia mangium and a number of other acacias produce good-quality timbers which are also well suited to the production of furniture and other higher-value products. A factor limiting this application is the occurrence of fungal heart rot in the central core of the stem at an early age. This defect does not seriously affect pulping properties if the crop is harvested early but it does significantly reduce the recovery of sawn wood and veneer from older logs grown for higher-value products. Identification of the factors causing the condition, and potential management strategies, have been the subjects of a major ACIAR project, ‘Heart rots in plantation hardwoods in Indonesia and southeast Australia’. Indonesian acacia plantations are also potentially threatened by root rot, another fungal disease. Root-rot diseases of A. mangium can cause crown dieback, reduced growth and tree death. In some areas, over 25 per cent of trees are affected. If not managed, this problem could seriously threaten Indonesia’s acacia plantation industry. ACIAR is now initiating a project to investigate it. A workshop with participants from Indonesia’s Forest Research and Development Agency, forestry companies and local universities, and research agencies in Australia and New Zealand, was conducted in Yogyakarta in February 2006 to review the results of the heart-rot project and define a strategy for root-rot research. These workshop proceedings provide valuable guidance on minimising the impacts of these two serious diseases in hardwood plantations.

Peter Core Director Australian Centre for International Agricultural Research

From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Contents

Foreword 3 Contents 5 Preface 7 Summary 8 Acacia mangium — a historical perspective on its cultivation Hardjono Arisman and Eko B. Hardiyanto 11 Hardwood plantation development and threats to its sustainability in Indonesia G.D. Golani 16 Heart rots in plantation hardwoods: the background to this ACIAR project Anto Rimbawanto 22 Heart rot and root rot in Acacia mangium: identification and assessment Caroline L. Mohammed, Karen M. Barry and Ragil S.B. Irianto 26 The mycology of the Basidiomycetes Ian A. Hood 34 The use of DNA techniques to identify fungi Morag Glen 46 Molecular identification of organisms associated with root and heart rot in Acacia mangium Morag Glen, Karina Potter and Purnamila Sulistyawati 55 Root rot in tree species other than Acacia A. Mohd Farid, S.S. Lee, Z. Maziah, H. Rosli and M. Norwati 60 The biological control of Ganoderma root rot by Trichoderma S.M. Widyastuti 67 Minimising disease incidence in Acacia in the nursery74 Budi Tjahjono 75 Developing a strategy for pruning and thinning Acacia mangium to increase wood value Chris Beadle 79 Options for solid-wood products from Acacia mangium plantations Eko B. Hardiyanto 87

Preface

About 1000 species of Acacia are native to Australia and neighbouring Indonesia, Papua New Guinea (PNG) and some Pacific island nations. Several are now used in commercial plantations in Southeast Asia and are the basis of substantial plantation- based industries. Within this region, plantations of tropical acacias have been established in China, Indonesia, Malaysia, Papua New Guinea, the Philippines, Thailand, Vietnam, and on Melville Island in Australia. The total area planted is now approaching 2 million ha and the largest of these estates (about 1.2 million ha) is located in Indonesia where the major species planted is Acacia mangium Willd. The majority of acacia plantations are used as feedstock for kraft pulp mills. However, substantial quantities of wood are now finding their way into markets based on high- value solid timber. Heart rot is caused by white-rot fungi which preferentially remove lignin. They are thought to enter trees through injuries and branch stubs. In Indonesia, this disease is a threat to solid-wood production and prevents the management of plantations for solid- wood values on some sites. Root rot is caused by basidiomycete fungi that can survive saprophytically on dead woody material or debris. The fungi attack the conducting tissues of living trees, rapidly causing tree death. The saprophytic nature of the fungi, together with the short rotations used by the pulpwood industry (6–7 years), have led to a rapid build-up of inoculum at some sites in Indonesia. Acacia mangium is particularly susceptible to root-rot disease on mineral soils in Sumatra and Kalimantan, where it is grown extensively. This volume brings together the current state of knowledge about these diseases. The more recent advances in knowledge have been made possible by funding through ACIAR project FST/2000/123 that supported a substantial body of research on heart rot. A project extension further encouraged the development of a knowledge base on root rot in Indonesia that will now be carried forward in a new ACIAR project, FST/2003/048. The research has been undertaken to date in close cooperation with the Indonesian Ministry of Forestry through its Centre of Plantation Forestry and Development, Gadjah Mada University’s Faculty of Forestry and PT. Musi Hutan Persada. The contributions of other Indonesian-based companies and the Forest Research Institute of Malaysia (FRIM) to the workshop ‘Heart rot and root rot in tropical Acacia plantations: a synthesis of research progress’ held in Yogyakarta on 7–9 February 2006, from which these proceedings are drawn, ensured a comprehensive coverage of the available information. The cooperating partners in Australia were the University of Tasmania and Ensis (the joint forces of CSIRO and Scion). The interest and expertise of the chapter authors is gratefully acknowledged.

7 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Summary

The total area planted to tropical acacias is now assessment of two trials within different commercial approaching 2 million ha. The largest of these estates growing areas in Indonesia (Riau and South Sumatra) (about 1.2 million ha) is in Indonesia where the major showed that provenance could influence the inci- species planted is Acacia mangium Willd. Acacia dence of heart rot. The basis for this observation mangium is generally grown as an exotic. It was first remains unclear, as heart-rot incidence was not cor- introduced into Malaysia in 1966 and into Indonesia related with the content of polyphenolic wood extrac- in 1979 where it was used in South Sumatra as a fire tives which are a known antifungal defence. break, for land rehabilitation and for reforestation of Root-rot diseases of A. mangium are associated alang-alang grassland. Its excellent adaptability to with crown dieback, reduced growth and tree death. degraded sites, rapid growth and wood properties When first detected, infected trees or disease foci quickly led to its commercial exploitation. In Indo- tend to be randomly distributed but then enlarge and nesia, pulpwood production from A. mangium plan- may aggregate. The rate of disease progress appears tations is currently more than 9 million m3/year, to be positively correlated with current levels of root while the potential for solid-wood production is rot. Accurate surveys to investigate spread are around 165,000 m3/year. Thus, domestication of the required and should record above-ground symptoms, species has progressed rapidly with extensive inspect the extent of root infection, and observe pat- research on its genetics, silviculture and wood utili- terns of disease infection. It is recognised that such sation. These proceedings add a new focus by pro- surveys are operationally laborious and costly. viding an insight into two diseases that threaten the Hence, survey options based on remote sensing sustainability of pulp and solid-wood industries should be considered. Root-rot diseases are by no based on tropical acacias. means isolated to acacias, and surveys to identify the The sustainability of planted forests in the tropics disease organisms are being conducted in forest plan- is threatened by their improper management, fire, tations of Azadirachta excelsa, Tectona grandis and and the illegal logging of native forests. Combating Khaya ivorensis throughout Peninsular Malaysia. these threats is a challenge that should be met through Several significant root diseases that affect planta- effective forest management and a multi-stakeholder tions in tropical Asia are caused by certain species of approach: the latter, in particular, is essential in the basidiomycetes. This group of fungi produces sexual drive against illegal logging. Sustainable manage- spores on the outside of microscopic structures called ment therefore embraces the interests of the planet – basidia which are held on macroscopic fruit bodies conserving biodiversity and preventing environ- such as mushrooms, toadstools, puff balls, earthstars, mental degradation; the people – providing opportu- and bracket, shelf, crust and coral fungi. Basidiomyc- nities for social development and poverty alleviation; etes occupy many niches, e.g. decomposing litter, and profit – ensuring a steady supply of renewable, decaying wood and soil organic matter, in a variety of high-quality, internationally cost-effective fibre. habitats that includes forests. Some species form ben- These interests are all threatened if monocultures are eficial mycorrhizal relationships with the roots of not managed to minimise the effects of diseases on host trees, but others are pathogens of the foliage, sustained production across rotations. stems or root systems of many tree species, including Heart rot is a major disease problem in A. man- acacias. Basidiomycete species are traditionally gium, particularly where trees are grown for solid- identified by the form and microscopic structure of wood products. Heart-rot fungi are wound parasites their fruitbodies, and by their appearance when iso- that enter through broken branches and branch stubs lated in laboratory culture. Important root diseases of after self pruning, singling and manual pruning. The trees in Indonesia are caused by the basidiomycete fungi attack cellulose and lignin, causing a typical fungi Rigidoporus microporus, Junghuhnia vincta, white rot that is associated with changes in colour, Phellinus noxius, and certain species of Ganoderma. texture and quality of the wood. These changes have In Peninsular Malaysia, Rigidoporus lignosus and been used to rapidly assess the incidence and severity P. noxius are the two major root diseases of A. of heart rot on harvested log-ends in the field. An excelsa, T. grandis and K. ivorensis. Root rots are

8 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. characterised according to the colour of the infected tions, though Trichoderma spp. have been shown to fungal tissues/roots. DNA-based molecular methods act as biological control agents of Ganoderma. have identified Ganoderma philippii as the causative Biological control of plant pathogens aims to agent of red root rot in A. mangium. This disease is reduce dependence on chemical treatments that may considered a major threat to the efficiency of wood cause environmental pollution and the development production by the pulpwood industry in Indonesia. of resistant strains. Filamentous fungi such as Tri- Traditional taxonomic methods for describing choderma are mycoparasites of plant pathogens and fungi causing heart rot and root rot continue to play an thus have potential for the biocontrol of plant disease: important role in identifying the causal agents of these species of Trichoderma are among the most widely diseases. However, molecular techniques for fungal tested agents. Although the mechanism of mycopara- identification are gaining in popularity, value and sitism is not fully understood, expression of extracel- effectiveness. DNA provides an abundance of taxo- lular cell-wall degrading enzymes is assumed to be nomic characters for the identification of organisms involved, including the action of chitinolytic and glu- that have inadequate morphological characters, or canolytic enzymes. As reported for other chitinolytic possess distinguishing features only during particular systems, endochitinase (EC 3.2.1.14) is among the stages of their life cycle. At present, because of their most effective for both antifungal and lytic activities speed, sensitivity and high throughput, variations of in comparison with other types of chitinolytic PCR-based techniques are popular for assessing these enzymes. Recently, a 32-kDa endochitinolytic DNA characteristics. Nevertheless, the exact method enzyme has been purified from Trichoderma reesei. selected for a particular application will depend on These Trichoderma isolates have an antagonistic several factors, including the number of samples and ability against some plant pathogenic fungi, such as number of candidate species. For heart rot and root Ganoderma spp., R. microporus, Rhizoctonia spp., rot, large numbers of fungal species are implicated, so Fusarium spp., and Sclerotium rolfsii and can effec- a technique that can simultaneously identify many tively suppress the development of these fungal path- species is more efficient than one based on species- ogens in vitro and in glasshouse experiments. specific probes or primers. In addition, as no compre- Disease management is not just an issue out in the hensive database of root- and heart-rot fungi from plantations. The foliar diseases Pestalotiopsis leaf A. mangium exists, techniques that require an exact spot, Phaeotrichoconis leaf spot, bacterial leaf blight match to a known species will provide only limited caused by Xanthomonas , phyllode rust disease information. All DNA-based techniques depend on an caused by Atelocauda digitata, and anthracnose adequate herbarium source of carefully prepared disease and tip necrosis caused by Colletotrichum sp. specimens with detailed morphological descriptions are all associated with tropical Acacia seedlings to verify the DNA-based protocol. being raised in nurseries, as are Pythium, Rhizoc- Although a new challenge for growers of tropical tonia, and Fusarium fungi that commonly cause acacia plantations, root-rot diseases have a long damping-off. Disease control requires integrated history of association with forest monocultures. This mnagement strategies based on a detailed knowledge has inevitably attracted research initiatives to under- of these pathogens and their interactions with the stand the dynamics of the diseases involved and seedlings and their environment, including the wider options for disease control. In Peninsular Malaysia, context of the nursery operation. poor land preparation and areas with a previous Acacia mangium wood has proved to be not only history of root disease were strongly associated with suitable for producing high-quality pulp and paper, the incidence of R. lignosus and P. noxius in forest but also an excellent material for solid-wood prod- plantations. Based on experience gained from the ucts. In Indonesia, there has been growing interest in management of root-rot disease in rubber plantations, utilising wood of A. mangium for solid wood, corre- good land management, the construction of isolation sponding with the declining availability of logs from trenches and the application of fungicides are consid- native forests. When managed for pulpwood, ered valuable tools in the control of root-rot disease A. mangium plantations are established at around in forest tree plantations. To date, cost-effective man- 1000 stems/ha and are clear-felled at age 6–7 years. agement tools based on biological and chemical Silvicultural techniques that incorporate thinning and treatments to soil and stumps have yet to be devel- pruning from below are of crucial importance in oped for controlling root rot in A. mangium planta- growing plantations for solid wood, as there is poten-

9 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. tial for large and persistent branches to develop in species that appears very susceptible to heart rot and, plantations: both are associated with the develop- if acacia wood is to be grown on high-risk sites, it ment of heart rot. Thinning systems must ensure that may be necessary to consider alternative species or final-crop trees retain green branches until pruning is hybrids that demonstrate an inherent resistance to completed as well as maintaining acceptable rates of heart rot. This need to focus on the host rather than growth of the retained trees. Form-pruning — the the pathogen to manage the disease makes even more selective removal of large branches or those com- sense now that it has been established that a suite of peting with the leader — ahead of lift-pruning, has fungi causes heart rot. been shown to increase the straightness of trees in a We did not need a workshop to come to the conclu- silvicultural system based on a final crop of about sion that root rot is a more intractable problem than 300 stems/ha pruned to 4.5 m. Acacia mangium can heart rot. Not only does root rot kill A. mangium, other be grown for solid-wood products with a rotation of organisms that fall into the same basket of diseases around 10 years, which is expected to produce a total kill trees across a range of species in the temperate as stem volume of more than 200 m3 per ha: about 30% well as the tropical zone. Solutions have been difficult of it will be for solid wood. The minimum tree diam- to find: to date, biological control has been shown to eter at breast height will be 30 cm. Heart rot is exac- work effectively for only one root-rot disease, that erbated by pruning when the plant material is caused by Heterobasidion annosum. If there is a chal- susceptible and a sufficient source of fungal inoculum is present to invade pruning wounds. lenge here, it is to integrate the breadth of skills and resources that are available in both the private and The workshop from which these proceedings are derived was billed as ‘a synthesis of research public sectors in the region to at least understand progress’: what have we learnt about heart rot and disease behaviour and then do the right things on the root rot to date and what are the challenges that must ground to contain the disease and, if possible, begin be confronted? moving towards disease management based on bio- The incidence of heart rot will probably exclude logical control. The heart-rot project demonstrated it the sustainable production of A. mangium for its was possible through research to make progress by solid-wood values on some sites. While we now have embracing a multi-stakeholder approach on how to a quick way of assessing the incidence and severity of manage a more tractable disease. Are we now ready to heart rot that will help determine disease risk, what meet the more difficult challenge? defines a high-risk site remains unknown. As with Caroline Mohammed other tree crops, good silviculture that includes Anto Rimbawanto pruning prescriptions based on live-branch pruning Joint Project Leaders should reduce disease incidence. Use of selected lines and the best seed source — one producing ACIAR Project FST2000/123 straight-growing and small-branched trees, may also ‘Heart rot in plantation hardwoods in contribute to managing disease incidence. However, Indonesia and south-east Australia’ in the absence of better information, A. mangium is a

10 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Acacia mangium — a historical perspective on its cultivation

Hardjono Arisman1 and Eko B. Hardiyanto1,2

Abstract The paper describes the cultivation of Acacia mangium from a historical perspective. The first introduction of the species to Sabah (Malaysia) in 1966 and South Sumatra in 1979 was for use as a fire break, land rehabilitation and reforestation of alang-alang (Imperata cylindrica) grassland. Eventually, A. mangium was planted in commercial plantations due to its excellent adaptability to degraded sites, rapid growth and good wood properties. In Indonesia the species has now become very important, with over 1 million ha planted in industrial forest plantations. Pulpwood production from A. mangium plantations is currently more than 9 million m3/year, while the potential for solid-wood production is around 165,000 m3/year. The domestication of the species has progressed rapidly, with extensive research in a range of aspects including genetics, silviculture and wood utilisation.

Acacia mangium Willd. has emerged as a key species provinces of Papua (Manokwari, Merauke) and grown for industrial forest plantations in Indonesia. Maluku (Seram, Aru). From a total of around 2.5 million ha of industrial plan- The first introduction of A. mangium for use as a tations, over 1 million ha have been established to plantation species was in Sabah, Malaysia in 1966 by A. mangium. The current use of A. mangium wood is D.I. Nicholson, an Australian forester. The seed was primarily for pulp and paper. Other uses include originally collected from a single tree at Mission medium-density fibreboard, furniture, plywood, Beach (Queensland). The initial seedlings were flooring and light construction. This paper presents a planted at Ulu Kukut as a firebreak. The species grew historical perspective of A. mangium cultivation. remarkably well and was subsequently used for reforestation of grasslands of alang-alang (the shade Domestication intolerant grass, Imperata cylindrica) and planted for commercial plantations in 1976 (Udarbe and Acacia mangium is native to northern Queensland, Hepburn 1986; Pinyopusarerk et al. 1993). Australia, mostly in the coastal tropical lowlands. Following its successful introduction to Sabah, The species also occurs naturally in the Western A. mangium was planted, and is now grown exten- Province of Papua New Guinea and the Indonesian sively, in many countries, including Indonesia, Thai- land, Vietnam and the Philippines, transforming 1 PT. Musi Hutan Persada, PTC Mall Blok I-9, Jl. R. what was once an unknown species with limited use Sukamto, Palembang 30114, Indonesia. to a popular tropical tree species (Pinyopusarerk et al. Email: . 1993). 2 Faculty of Forestry, Gadjah Mada University, Bulaksumur, Yogyakarta 55281, Indonesia. In Indonesia, the early introduction of A. mangium Email: ; . 1979 into South Sumatra. Seeds consisting of three

11 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. seedlots were brought by Mr R. Darmono (a former Following the successful species trial in Suban- director of the Seed Division, Directorate General of jeriji, seed from four provenances of Queensland’s Forestry) from Sabah (Malaysia) and given to Hard- Cairns region, namely Cassowary, Jullaten, jono Arisman for use in a field trial in Subanjeriji, Mossman and Daintree, were imported in 1980. South Sumatra. The object of the trial was to examine Provenance resource stands were established in Sub- the potential for using the species for the rehabilita- anjeriji (300 ha) and Benakat (100 ha) using these tion and reforestation of large areas of alang-alang new Australian provenances and are referred to as the grassland existing in the region. The plots of the Subanjeriji local landrace. Each provenance con- initial seedlots from Sabah still exist today and sisted of 10 individual parent trees. In 1982, seeds remain in fairly good condition (Figure 1). collected from Sidei (Manokwari) by E.B. Hardi- In South Sumatra, A. mangium grows reasonably yanto and Seram (Maluka) by M. Subagyono were well on former alang-alang grassland. The species has also planted in Subanjeriji. Seed from Sanga-Sanga a rapid initial growth rate and starts closing its canopy (East Kalimantan) was also introduced into Suban- early, suppressing the growth of alang-alang quite jeriji, but the growth of the Sidei, Seram and Sanga- effectively. In a species trial conducted in Subanjeriji Sanga provenances was poor. by the Directorate of Reforestation and Land Rehabil- The Subanjeriji provenance seed stands were used itation and a Japan International Cooperation Agency as seed production areas managed by PT. Inhutani I. project in the early 1980s, A. mangium emerged as the Seeds collected from these production areas were most promising in terms of adaptability and growth. used for reforestation programs and plantation devel- The site was dominated by red–yellow podsolic soil opment all over Indonesia during the late 1980s. Seed (Ultisol or Oxisol) that is inherently acid and poor in production in these areas ceased in 1995 when seed nutrient reserves. Acacia mangium, a leguminous spe- of better genetic quality introduced from other prov- cies, thus appears to adapt well to poor soils. enances became available.

Figure 1. The plot of the first introduction of Acacia mangium into Subanjeriji, South Sumatra established in 1979. This photograph was taken in January 2006.

12 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Availability of genetic materials by the government to develop 2.3 million ha of industrial forest plantations by 2000 and 10.5 million ha by In the early 1980s, information on the genetic varia- 2030 (Ginting et al. 1996). Only about 2.5 million ha bility of A. mangium was sparse and it was not known had been realised as of the end 2004, with which of the provenances should be used for planta- A. mangium the primary species planted. The proper- tion development. Indeed, seeds of many provenances ties of pulp and paper processed from A. mangium were not available until the CSIRO Division of Forest wood are known to be excellent and comparable with Research, Australia, with FAO support, made collec- or better than those of Eucalyptus species used for tions in the 1980s (Doran and Skelton 1980; Turnbull paper making. Acacia mangium pulp has fairly good et al. 1983). Another collection was undertaken in formation and opacity which is excellent for paper 1983 within the framework of the Cooperative Seed and tissue (Palokangas 1996) and the pulp has been Collection Program between the Office of Forests, accepted in the international pulp market and com- Papua New Guinea and FAO’s Forestry Department. petes reasonably well with other short fibre species. Seed of A. mangium from these collections, together The productivity of A. mangium plantations varies with seed collected by the Directorate-General of For- depending upon a number of factors including seed estry, Indonesia, was used for provenance trials in the quality, site characteristics and silvicultural practices. early 1990s in many parts of Indonesia. Current mean annual increments of total volume are The results of provenance trials indicated that seed in the range of 20–33 m3/ha, but can reach more than of the Subanjeriji local landrace and the Cairns 40 m3/ha/year on the best sites (Hardiyanto 2005). region of Queensland, Australia had suboptimal Research and development on A. mangium has growth, while Far North Queensland (FNQ), Papua been extensive across a range of fields, including tree New Guinea (PNG) and Muting Irian Jaya prove- breeding, silviculture, growth and yield, pests and nances grew better (Havmoller 1991; Hardiyanto et diseases, and wood and non-wood products. With al. 1994; Hardiyanto et al. 1997; Leksono and Ros- iawan 1997; Hardiyanto et al. 2000). The suboptimal regard to tree breeding, second-generation improve- growth of the Subanjeriji local landrace was not sur- ment programs are in progress. Studies on site man- prising since this seed source had originated from the agement and the productivity of the second rotation Cairns region and its genetic base was known to be have also been conducted and show that the growth narrow both at growth and molecular levels (Butcher rates can be maintained or even increased, provided et al. 1996, 1998). Renewed efforts to broaden the proper silvicultural practices are employed (Hardi- genetic base of A. mangium were undertaken in the yanto et al. 2003). early 1990s by introducing seeds from a large Various studies have been conducted to assess the number of provenances of PNG, FNQ and Muting. suitability of A. mangium wood for solid-wood prod- Collections were made by a number of institutions ucts such as furniture, flooring and plywood (Awang including CSIRO’s Australian Tree Seed Centre and Taylor 1993). A recent study on the mechanical (ATSC), the Department of Forestry, Faculty of For- properties of sawn timber indicated that the wood of estry, Gadjah Mada University, and several forestry A. mangium can be used for structural materials and companies and now form the basis of seed production meets the requirement of the Indonesian Standard for areas and seed orchards. Since the mid 1990s, Wood Construction (Amalia 2003; Firmanti and improved seed with a broader genetic base has Kawai 2005). The development of A. mangium plan- become generally available for operational planta- tations for solid-wood utilisation is in progress. tions (Hardiyanto 1998). Large-scale A. mangium plantations have been developed mainly by forestry companies. More Plantation development recently, however, smaller-scale plantations of A. mangium have been established in outgrower Acacia mangium has become the leading tree species schemes by individual farmers on the farmers’ land in forestry plantation programs in several Asian in cooperation with forestry companies which cover countries (Awang and Taylor 1993; Turnbull et al. the costs of establishment and maintenance: the ben- 1998). In Indonesia, the large-scale development of efits are shared between the farmers and companies. A. mangium plantations began in the early 1990s, Small-scale plantations may also be established by corresponding with an ambitious program launched farmers using their own capital.

13 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

As A. mangium is an exotic species being planted Southeast Asia. It is only 20 years since the first on a large scale, the potential threats to plantations introduction of this species to South Sumatra. In this have also been identified and studied. Heart rots have short time, the natural variation within the species has been reported to attack older trees, which can reduce been assessed, breeding programs established, timber volume and quality. Heart-rot surveys in molecular marker technologies applied, silvicultural Malaysia (where A. mangium plantations are estab- studies completed, growth and yield studied, and lished for the production of sawn timber) reported wood and fibre properties determined. Such rapid that heart-rot incidence varied from low to high. This progress in domesticating this tree species is remark- caused the Malaysian Government to review its able. Nonetheless, research in these fields needs to policy on the further development of A. mangium continue in order to improve plantation productivity plantations in Peninsular Malaysia (Lee et al. 1996). and sustainability. Environmental and biodiversity A recent survey at a number of sites in Indonesia aspects of A. mangium plantations also require revealed that heart-rot incidence differed between further study. regions, ranging from low to high, depending upon a combination of climate, plantation management References (pruning) and age. In regions in which heart-rot incidence was low, such as in South Sumatra and East Amalia, L. 2003. Pengujian mutu kayu Acacia mangium Kalimantan, the development of sawlog plantations dengan menggunakan alat Panter. Publikasi Divisi R&D was considered feasible (Barry et al. 2004). PT Musi Hutan Persada, Technical Notes 13(5). Another disease that has been causing concern in Awang, K. and Taylor, D., ed. 1993. Acacia mangium, A. mangium plantations is root-rot, associated with growing and utilization. FAO, Bangkok, Thailand, and the fungi Ganoderma sp. and Rigidoporus lignosus. Winrock International, MPTS Monograph Series No. 3, The disease often occurs in patches with a concentric 280p. pattern of spread. Trees attacked by the disease show Barry, K.M., Irianto, R.S.B., Santoso, E., Turjaman, M., crown thinning and subsequent death. Research on Widyati, E., Sitepu, I. and Mohammed, C.L. 2004. Incidence of heartrot in harvest-age Acacia mangium in this problem is continuing in a number of projects. Indonesia using a rapid survey method. Forest Ecology and Management, 190, 273–280. Wood production Butcher, P.A., Moran, G.F. and Perkins, H.D. 1996. Genetic resources and domestication of Acacia mangium. In: Wood production from A. mangium plantations was Dieters, M.J., Matheson, A.C., Nikles, D.G., Harwood, started in the early 1990s. The wood was used mainly C.E. and Wakler, S.M., ed., Tree improvement for for pulp and medium-density fibreboard. Initially, sustainable tropical forestry. Proceedings of QFRI– IUFRO Conference, Caloundra, Queensland, Australia, the volume of wood produced was relatively small, 467–471. increasing markedly in the year 2000. Wood produc- — 1998. RFLP diversity in the nuclear genome of Acacia tion in Indonesia for pulp and medium-density fibre- mangium. Heredity, 81, 205–213. board harvested from A. mangium plantations is Doran, J.C. and Skelton, D.J. 1980. Seed collection of roughly as follows: Riau and Jambi, 5,860,000 m3/ Acacia mangium for international provenance trial. 3 year; South Sumatra and Lampung, 2,500,000 m / Forest Genetic Resources No. 11. 3 year; East and South Kalimantan, 750,000 m /year; Firamanti, A. and Kawai, S., 2005. A series of studies on the 3 West Kalimantan 200,000 m /year. The solid-wood utilization of Acacia mangium timber as structural utilisation of A. mangium is increasing with potential materials. Proceedings of the 6th International Wood supply, which in Indonesia is currently estimated at Science Symposium, LIPI–JSPS Core University 165,000 m3/year (Thorp 2005). Program in the Field of Wood Science: Towards ecology and economy harmonization of tropical forest resources, Bali, 463–473. The way forward Ginting, N.A., Daryono, H. and Siregar, C.A. 1996. Ecological aspects of forest plantation. In: Otsamo, A., Over the past 25 years, A. mangium has gone from a Kuusipalo, J. and Jaskari, H., ed., Proceedings of virtually unknown tree in the wilds of North Queens- workshop on reforestation: meeting the future industrial land, Papua New Guinea and Irian Jaya to a major wood demand, 20 April–1 May 1996, Jakarta, Enso commercial plantation species for pulp and paper in Forest Development Oy Ltd.

14 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Hardiyanto, E.B. 1998. Approaches to breeding acacias for IUFRO symposium on the impact of diseases and insect growth and form: the experiences at PT. Musi Hutan pests in tropical forest, 1–10. Persada. In: Turnbull, J.W., Crompton, H.R. and Leksono, B. and Rosiawan, H. 1997. Evaluasi uji provenansi Pinyopusarerk, K., ed., Proceedings of an international Acacia mangium umur 30 bulan di Kampar Riau. Bulletin workshop held in Hanoi, Vietnam, 27–30 October 1997, Kehutanan, 32, 15–22. 178–183. Palokangas, E. 1996. Acacia mangium in pulp and paper — 2005. Beberapa isu silvikultur dalam pengembangan production. In: Otsamo, A., Kuusipalo, J. and Jaskari, H., hutan tanaman. Dalam: Hardiyanto, E.B., ed., Prosiding ed., Proceedings of workshop on reforestation: meeting Seminar Nasional Peningkatan Produktivitas Hutan- the future industrial wood demand, 20 April–1 May 1996, Peran Konservasi Sumberdaya Genetik, Pemuliaan dan Jakarta, Enso Forest Development Oy Ltd. Silvikultur dalam Mendukung Rehabilitasi Hutan. ITTO Pinyopusarerk, K., Liang, S.B. and Gunn, B.V. 1993. dan Fakultas Kehutanan UGM, Yogyakarta, 26–27 Mei Taxonomy, distribution, biology and use as an exotic. In: 2005.hal.121–134. Awang, K. and Taylor, D., ed., Acacia mangium, growing Hardiyanto, E.B., Siregar, S.T. and Chandralika, 1997. Hasil and utilization. FAO, Bangkok, Thailand, and Winrock uji provenans Acacia mangium pada umur 28 bulan di International, MPTS Monograph Series No. 3, 1–20. Setuntung. Dep. R&D PT. Musi Hutan Persada, Thorp, A. 2005. Sources of furniture grade acacia timber Technical Notes, 3(4). from plantations in Indonesia. Final report prepared for the International Finance Corporation, Programme for Hardiyanto, E.B., Siregar, S.T., Wahyono, R., and Rokhim, Eastern Indonesia SME Assistance. M. 2000. Hasil uji provenans Acacia mangium umur 5,5 tahun di Setuntung. Dep. R&D PT. Musi Hutan Persada, Turnbull, J.W., Crompton, H.R. and Pinyopusarerk, K., ed. 1998. Recent developments in Acacia planting. Technical Notes, 10(15). Proceedings of an international workshop held in Hanoi, Hardiyanto, E.B., Sutarjo and Simanjuntak, S. 1994. Vietnam, 27–30 October 1997. Canberra, ACIAR Beberapa provenans Acacia mangium introduksi baru Proceedings No. 82, 383p. tumbuh lebih baik dibanding seedlot lokal. Publikasi Turnbull, J.W., Skelton, D.J., Subagyono, M. and R&D PT. Musi Hutan Persada, Technical Notes, 4(3), 4p. Hardiyanto, E.B. 1983. Seed collections of tropical Havmoller, P., Wahyuningrini, S., Nurjanah, N. and acacias in Indonesia, Papua New Guinea and Australia. Indriany, M. 1991. Seven years growth of acacia species Forest Genetic Resources Information, 12, 2–15. and provenance trial at Subanjeriji, South Sumatra. Udarbe, M.P. and Hepburn, A.J. 1986. Development of Acacia Publikasi R&D PT. Musi Hutan Persada, Technical mangium as plantation species in Sabah. In: Turnbull, J.W., Notes, 1(1), 5p. ed., Australian acacias in developing countries. proceedings Lee, S.S., Ibrahim, Z., Noor, H.M. and Wan Razali Wan of international workshop held at the Forestry Training Mohd 1996. Impact of heart rot in Acacia mangium Willd. Centre, Gympie, Queensland, Australia. Canberra, ACIAR plantations of Peninsular Malaysia. Proceedings of Proceedings No. 16, 157–159.

15 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Hardwood plantation development and threats to its sustainability in Indonesia

G.D. Golani1

Abstract

Sustainable management of plantation forests should embrace the interests of the planet — conserving biodiversity and preventing environmental degradation; the people — providing opportunities for social development and poverty alleviation; and profit — ensuring a steady supply of renewable, high quality, internationally cost-effective fibre. Options for managing plantations in this way are considered. The sustainability of plantation forests is threatened by improper management of monocultures, fire and illegal logging. This paper outlines ways of combating these threats through effective forest management and a multi- stakeholder approach to ending illegal logging. Interdisciplinary research will continue to play a role in the current and future establishment of industrial forests in Indonesia.

Indonesia is one of the major players in pulp and paper natural forest will eventually cease: natural forests will production. Strong demand for paper in Asia led to an be preserved and valued for their ecological functions overall increase in the global pulp market by 2.9% in and benefits. To fulfil this requirement, development of 2003 (APRIL 2004). Strategic location, proximity to new forest plantations in balanced and responsible the fast-developing economic giants, China and India, ways is of critical importance. This paper is intended to and climatic and physiographic conditions conducive discuss the development of industrial plantation forests for growing short-rotation fibre plantations make Indo- in Indonesia and constraints to their sustainability. nesia a preferred choice for pulp and paper production. In line with ever-increasing demands for wood, Indo- Key issues nesia has initiated a massive reforestation program. Since the mid 1980s the area of industrial forest plan- The main issue in the management of large-scale tations in Indonesia, particularly those of short-rotation industrial plantation forests of fast-growing species species, has increased dramatically with a target of 10.5 is to ensure product sustainability over successive million ha by 2030 (Gintings et al. 1996). The aim of rotations. This is particularly true in areas conducive the program is to provide the required supply of forest to high growth rates and thus imposing heavy products while at the same time maintaining the demands on site resources. Nambiar and Brown remaining natural forest. It is expected that, after 2020, (1997) have set goals for industrial plantation for- most of the forestry industry in Indonesia will depend estry, which include: more on wood from plantation forest than natural forest • productivity — to ensure that the trend in (Natadiwirya 1998). Dependence upon exploitation of plantation productivity is non-declining over successive harvests 1 PT. Riau Andalan Pulp and Paper, PO Box 1089, • soil and water conservation — to protect and if Pekanbaru 28000, Indonesia. possible enhance the quality of soil and water in Email: . the plantation environment,

16 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

• economic viability — to promote incentive, mercial, environmental and social development. Pro- innovation and profit for the business of growing tection and conservation of HCVF (high and utilising wood conservation value forest), riparian areas to protect • socioeconomic improvement — to improve the soil and maintain natural river ecosystems, and buffer economic and social benefits to the community. zones to serve as habitat and corridors for wildlife, Sustainable management of plantation forest is not form part of this approach to maintain the balance merely about maintaining tree growth, rotation after between plantations and natural forests for sustain- rotation. Rather, it includes a more holistic approach able outcomes. that fosters the long-term sustainability of the entire Social aspect is another important component of forest ecosystem and considers the diverse social plantation forest management. In the past this aspect impacts. Forest management should recognise and received relatively little consideration: the main involve a consideration of several values, especially emphasis was wood production (Gintings and those of economic, environmental, and social signif- Suharti 1998). Social problems such as conflicts icance. Plantation forest should be mutually accept- between companies and local communities over able to various stakeholders: access to land, and years of experience, have indi- • to industries, for steady sustained wood supply cated the importance of a participatory process in • to local people who need access to land and forest plantation forest management. The evidence sug- resources, minor forest products like firewood, gests that such a process contributes to sustained honey, herbs etc. forest productivity as well as the welfare of forest • to governments and environmental groups for communities. Thus, active participation of local conservation of natural forests and the ecosystem. people should be continuously encouraged. The interactions between people, forests and other natural resources have to be better understood. Man- Challenges agement of plantation forest should be considered as a business that is efficient and economically benefi- There has been growing concern over the sustaina- cial for all the above stakeholders including conces- bility of wood production in industrial plantation sionaires and local people by providing opportunities forest management systems (Muhtaman et al. for employment. This will also make plantation 2000). Without proper management, long-term site forests not only viable and sustainable but attractive productivity may fall. This is particularly relevant to investors. Inefficient management systems may to industrial plantation forest in Indonesia; i.e. a cul- create potential social problems that lead to rapid tivation system involving monocultures of fast- conversion of forest resources to other land uses. growing species, many of them exotic. Under this Parthama and Widiarti (1998) emphasise that man- system, improper management negatively affects agement options for plantation forest, low or high the hydrological balance of river basins within the capital, must be carefully considered. plantation forest areas, soil nutrients, and the food Ecological aspects of plantation forest establish- chain (Santosa 1998), leading to decreased quality ment are critical. These should include conservation of soil and water as well as higher vulnerability to of environment and biodiversity to maintain the eco- potential pests and diseases. Forest fires and illegal logical functions of the forest landscape. Thus, logging also constrain the sustainable production of changes to the environment during plantation estab- industrial plantation forest in Indonesia. Manage- lishment should be minimised. Strengthening of the ment options that simultaneously meet wood-pro- relationship between production forests and natural duction goals and avoid these threats should be landscapes includes the retention of conservation developed. areas in such a way that losses to biodiversity are also Plantation forests are often established on mar- minimised (Santosa 1998). Failure to recognise this ginal soils. One constraint to the further expansion of requirement in plantation forest management may the plantation forest, especially of fast-growing spe- deleteriously affect the long-term sustainability of cies, is long-term site productivity. Each harvest is the forest landscape. accompanied not only by nutrient export, but also The ‘mosaic’ plantation concept (Figure 1) offers some soil erosion, soil compaction, nutrient leaching an important strategy in managing forest resources in and volatilisation. Losses of nutrients during harvest a sustainable manner that balances the need for com- and site preparation may exceed the rate of their

17 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. replacement by weathering of minerals in soils or by the availability and use of good genetic stock. Tree nutrient inputs through precipitation (Ruhiyat 1998; improvement programs are therefore crucial to the Santosa 1998). Thus, a proper fertiliser regime and adoption of monoculture plantations. Wood volume effective soil and water conservation strategies are and density are among the important characteristics of crucial to sustainable plantation forestry. interest. Through intensive breeding programs, Wong Fertiliser application can replenish the nutrient and Wijoyo (2005) have shown that predicted genetic supply to maintain or even increase productivity. The gain in A. mangium can increase to as high as 43% for results of Riaufiber research in Baserah, Riau, indi- wood volume and 5% for wood density, with pre- cate that application of balanced NPK fertilisers and dicted mean annual increments (m3/ha/yr) of 66.0 good harvest-residue management lead to increased (potential) and 46.2 (operational) (Table 1). In addi- height and diameter at breast height of 3-year-old tion to wood volume and density, Riaufiber breeding Acacia mangium in the second compared with the programs also consider resistance to pests and dis- first rotation at the same age (Siregar et al. 2004). eases. Through selection, Riaufiber has demonstrated This improved growth was in part attributable to that some Acacia crassicarpa plus-trees have genetic better genetic material. Attention to detail at this level resistance to Passalora (formerly Pseudocercospora) should mean that degraded forest lands can also be leaf and shoot blight disease (Gafur et al. 2005). replanted and managed to provide wood, at the same The common use of a small number of species of time restoring ecosystem services such as freshwater the same age grown in monoculture in plantation regulation and soil retention. forest poses a concern for managers, as these condi- Sustainable production of industrial plantation tions provide an opportunity for pests and diseases to forest will also depend on the ability of managers to multiply and reach epidemic proportions (Sitepu and genetically improve planting stock. The more recent Suharti 1998). An abundance of substrate over a rel- industrial plantation forests in Indonesia have been atively long time provides particularly suitable con- established to mainly exotic species. This has limited ditions for the development of pests and pathogens.

Figure 1. Mosaic plantation concept in Raiufiber Industrial fibre plantations (HTI)

18 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. a (2 best) CP orchard Vol. Density Index (5 best) trees No. of /ha/year) – operational discount 3 families OP orchard CP orchard CP orchard Vol. Density Index No. of trees (2 best) No. of No. CP orchard families /ha/year) Predicted MAI (m 3 (5 best) /ha/year 3 Vol. Density Index No. of trees No. of Selection Predicted gain (%) Selection Predicted gain (%) Selection Predicted gain (%) No. of OP orchard CP orchard CP orchard families trees No. of No. 160 13,824 37 52 29.0 4.9 14.8 4 6 37.5 5.6 23.2 3 4 42.7 5.0 27.2 No. of families Superior 58.1 61.9 64.2 66.0 40.6 43.3 45.0 46.2 Predicted gain and mean annual increment (MAI) of selected trees in some Riaufiber AMPFT trials. OP is open-pollinated; CP close-pollinated. Operational discount rate = 30%. Test mean MAI ~ 45 m Trial code Tested OP orchard CP orchard CP orchard (5 best) AMPFT01AMPFT02 and AMPFT03 85 4320Trial code 12 PYI Category 21 27.7 0.3 26.4 3 Predicted MAI (m 4 38.6 0.4 37.6 1 1 41.1 0.8 43.7 AMPFT01AMPFT02 and SuperiorAMPFT03 a 57.5 62.4 63.5 40.2 43.7 44.4 Table 1.

19 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

One good example is root-rot disease. This disease, • organised large-scale logging in natural forest which is caused by Ganoderma sp., has been dam- without a permit aging oil-palm plantations in Southeast Asia for • smallholder farming — clearing of forest land for years. In acacia plantations, the disease is also agriculture without permit causing high mortality (3.2–40.0%) in Southeast • excessive logging of forest concessions Asia and India (Eyles et al. 2004). Although often • misuse of logging licenses — felling of trees in difficult in practice, the implementation of compat- other than designated areas. ible control measures, more popularly known as inte- To maintain the already very limited conservation grated pest management (IPM), to maintain pest and area (15–20% of concession area), forest managers disease losses at acceptable levels, is recommended. must provide alternative ways to plan and manage the This will include appropriate application of chemi- corridors of native forest that allow local people to cals, site- and species-specific silvicultural practices, collect firewood, medicinal plants and other products biocontrol, and planting of less-susceptible species. to support their livelihood. The management of plantation forest is a key factor Plantation forestry generates 5 direct and 30 indi- in pest and disease management: very often control rect jobs for each 100 ha managed and thus offers an measures fail due to improper management. option for a legitimate livelihood to people currently Forest fires cause devastating effects on the lands. engaged in illegal logging. A multi-stakeholder task In addition to the loss of thousands of hectares of force, involving local communities, law-enforcement wood, both in natural and plantation forests, fire also authorities, environment protection agencies, non- causes starvation or even extinction of wildlife. The government organisations and forest industries can smoke causes health problems in forest communities. be effective in combating illegal logging. A number of causal factors have been associated with forest fire in Indonesia, including ‘manual slash and burn’ land-clearing practices mainly adopted by Conclusions small farmers, escaped cooking fires, careless dis- The development of plantation forests in Indonesia is posal of cigarette butts and arson. Some farmers taking place and seen as part of future progress. For intentionally use fire to open up new land. Past sustainable production, plantation forests should be logging has also left forests degraded and very sus- properly managed by considering several values, ceptible to fire. Fire often gets out of control. The especially those that are of economic, environmental issue of fire can be solved only by prevention and and social significance. There is growing demand for effective management. Plantation forest manage- more-integrated approaches to natural resource man- ment should incorporate: agement. The interactions between all the resources • a no-burn policy for land clearing. (soil, water, biological and atmospheric resources) • plantation forest and wildfire fighting capability within a given landscape need to be better understood. • compulsory basic fire-fighting and prevention There will of course be more challenges ahead. These training for staff include quality of soil and water, plant genetic mate- • community awareness and participation in fire rial, vulnerability to pests and diseases, forest fire, and prevention and control. illegal logging. Careful consideration of these aspects • stronger regional cooperation in managing forest will help drive plantation forestry along the right fires will also help. track. Interdisciplinary research will continue to play Illegal logging has been occurring for years and is an important role in the current and future establish- believed to have destroyed millions of hectares of ment of industrial forests in Indonesia. forest. Wood is routinely smuggled between neighbouring islands and countries, costing the Indonesian References Government millions of dollars in loss of revenues each year. Illegal loggers also cut trees in the conser- APRIL (Asia Pacific Resources International Holding vation areas and in corridors left by plantation forest Limited) 2004. Sustainability report, 2004. Jakarta, managers for biodiversity conservation, causing Indonesia, APRIL. wildlife habitats to degrade. Illegal logging in Indo- — 2005. ‘Fact sheet’ on illegal logging –June 2005. Jakarta, nesia occurs in a number of ways (APRIL 2005): Indonesia, APRIL.

20 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Eyles, A., Mohammed, C., Barry, K. and Griffin, R. 2004. Harwood, C.E. and Booth, T.H., ed., Sustained Current and future management strategies of root rot productivity of short and medium rotation plantation pathogens in tree crops: critical literature review. forests for commercial and community benefit in Unpublished paper. Indonesia: an analysis of research priorities. Canberra, Gafur, A., Tjahjono, B., Wong, C.Y. and Wijoyo, F.S. 2005. CSIRO Forestry and Forest Products, 65–67. Evaluation of Pseudocercospora resistance of selected Ruhiyat, D. 1998. Management of productive capacity and Acacia crassicarpa clones. Unpublished report. soil improvement. In: Nambiar, E.K.S., Gintings, A.N., Gintings, N.A., Daryono, H. and Siregar, C.A. 1996. Ruhiyat, D., Natadiwirya, M., Harwood, C.E. and Booth, Ecological aspects of forest plantation. In: Otsamo, A., T.H., ed., Sustained productivity of short and medium Kuusipalo, J. and Jaskari, H., ed., Proceedings of a rotation plantation forests for commercial and workshop on reforestation: meeting the future industrial community benefit in Indonesia: an analysis of research wood demand, 30 April–1 May 1996. Jakarta, Enso priorities. Canberra, CSIRO Forestry and Forest Forest Development Oy Ltd. Products, 17–22. Gintings, N.A. and Suharti, S., 1998. Social factors, local Santosa, Y., 1998. Off-site impacts and threats to impacts and community interactions in timber estate environmental conditions including biodiversity. In: establishment. In: Nambiar, E.K.S., Gintings, A.N., Nambiar, E.K.S., Gintings, A.N., Ruhiyat, D., Ruhiyat, D., Natadiwirya, M., Harwood, C.E. and Booth, Natadiwirya, M., Harwood, C.E. and Booth, T.H., ed., T.H., ed., Sustained productivity of short and medium Sustained productivity of short and medium rotation rotation plantation forests for commercial and plantation forests for commercial and community benefit community benefit in Indonesia: an analysis of research in Indonesia: an analysis of research priorities. Canberra, priorities. Canberra, CSIRO Forestry and Forest CSIRO Forestry and Forest Products, 53–58. Products, 59–64. Siregar, S.T.H., Nurwahyudi and Mulawarman 2004. Muhtaman, D.R., Siregar, C.A. and Hopmans, P. 2000. Criteria and indicators for sustainable plantation forestry Growth and site productivity of a three-year-old Acacia in Indonesia. Bogor, Indonesia, CIFOR, 72p. mangium plantation from different harvesting residue management in Riau, Sumatera, Indonesia. Paper Nambiar, E.K.S. and Brown, A.G. 1997. Management of presented at the workshop on CIFOR network on site soil, nutrients and water in tropical plantation forests. Canberra, ACIAR Monograph, No. 43. management and productivity in tropical plantation forests, 22–26 November 2004, Sao Paolo, Brazil. Natadiwirya, M. 1998. Plantation forest in Indonesia: basic resource issues and national goals. In: Nambiar, E.K.S., Sitepu, I.R. and Suharti, M., 1998. Pest and disease Gintings, A.N., Ruhiyat, D., Natadiwirya, M., Harwood, management in industrial forest plantations in Indonesia. C.E. and Booth, T.H., ed., Sustained productivity of short In: Nambiar, E.K.S., Gintings, A.N., Ruhiyat, D., and medium rotation plantation forests for commercial Natadiwirya, M., Harwood, C.E. and Booth, T.H., ed., and community benefit in Indonesia: an analysis of Sustained productivity of short and medium rotation research priorities. Canberra, CSIRO Forestry and Forest plantation forests for commercial and community benefit Products, 13–16. in Indonesia: an analysis of research priorities. Canberra, Parthama, I.B.P. and Widiarti, A. 1998. Options for CSIRO Forestry and Forest Products, 39–48. management: low capital–high capital. In: Nambiar, Wong, C.Y. and Wijoyo, F.S. 2005. Predicted gain of E.K.S., Gintings, A.N., Ruhiyat, D., Natadiwirya, M., selected trees in AMFPT 01 to 03. Unpublished report.

21 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Heart rots in plantation hardwoods: the background to this ACIAR project

Anto Rimbawanto1

Abstract

Heart-rot fungi are wound parasites that enter through broken branches and branch stubs after self pruning, or through singling and manual pruning. Heart rot has been identified as one of the major disease problems in Acacia mangium. It was therefore important to address this problem if the benefits of tree improvement programs are to be realised, particularly where trees are grown for solid-wood products. The focus of the project activities was to assess the incidence of heart rots, including links with silvicultural practice and identify any resistance to disease. DNA-based methods were to be developed and coupled with traditional taxonomic methods to identify fungi causing heart rot. Technology-transfer activities included assisting plantation managers to reduce heart rot, and staff exchange and interaction. Preliminary work was also undertaken in respect of root rot for some of the above activities.

During the past decade, Australian and Southeast In Indonesia the demand for wood, from both Asian researchers have completed a number of collab- domestic and international markets, continues to orative forest pathology projects. Pathology and tree increase, and the Government of Indonesia has improvement specialists from CSIRO have collabo- embarked on a massive afforestation and reforesta- rated with their counterparts from government tion program aimed at preserving the natural forest research agencies and universities in Indonesia and while maintaining the supply of forest products. five other Southeast Asian countries with ACIAR and Since the mid-1980s, the area of forest plantations in Center for International Forestry Research (CIFOR) Indonesia, particularly those grown on short rota- support. One of the most successful of these projects tions, has dramatically increased. The government was the survey of diseases of tropical acacias carried has launched an ambitious program for rehabilitation out during 1995–96, which included a workshop in of unproductive Imperata grassland and secondary Subanjeriji. One request of the representatives at the scrubland into industrial forests in islands other than workshop was met by the publication of the book Java. The program aimed to establish 2.3 million ha ‘A manual of diseases of tropical acacias in Australia, of plantations by the end of 2000 and 10.5 million by South East Asia and India’ (Old et al. 2000). One of 2030. The current plantation program has been the major disease problems discussed at the workshop planned to produce wood for pulp, paper-making or was heart rot in Acacia mangium Willd. medium-density fibreboard. The Ministry of For- estry has identified the supply of genetically improved material as being critical to the success of 1 Ministry of Forestry FORDA, Centre for Plantation the plantation program. The genetic material being Forest Research and Development, Jalan Palagan Tentara generated is potentially significant in producing Pelajar Km. 15, Purwobinangun – Pakem, Yogyakarta high-quality plantation for solid-wood products and 55585, Indonesia. Email: . for achieving sustainability by optimising yield.

22 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Fast-growing hardwood plantations are important to stem, size and season of injury, age of wounds, stand the economies of many Australasian countries, dynamics and decay organism(s). including Australia and Indonesia. The major planta- Heart-rot fungi are generally pan-tropical or cos- tion species in Indonesia is A. mangium, which appears mopolitan saprophytic wood-decay fungi or wound to be highly susceptible to heart rot. Unless the problem parasites. A range of hymenomycetes are known to of heart rot is overcome it is unlikely that the benefits of be associated with heart rot of acacias, but several tree improvement programs will translate into fungi associated with A. mangium have only recently increased productivity and economic gain. The finan- been identified. cial interests of small growers and agroforestry in Indo- As heart-rot fungi are wound parasites, lopping off nesia must also be considered. Tree breeders work branches with parangs or machetes should be closely with their counterparts in resource protection, avoided. Singling of multi-stemmed, fast-growing tree physiology, soil and silviculture to develop sus- trees creates large stem wounds that heal slowly and tainable production systems. This type of activity best are more prone to invasion by fungi. Wound protect- fits the role of a public research institution rather than ants are commercially available but their effective- the private sector, as such organisations have devel- ness on A. mangium has not been tested. This oped these interlinking contacts and are less hampered approach is also expensive and impractical for large- by issues of intellectual property. scale use in forest plantations. Quantitative information on the extent of heart rot in acacia plantations is lacking. In Peninsular Malaysia, a Heart rot volume loss of up to 17.5% of the merchantable timber of A. mangium has been reported as a result of heart rot Wood-decay fungi may be defined by the zone of (Zakaria et al. 1994). Although the disease is also the tree that the fungi invade. The outer structure of known to be present in A. mangium plantations in Indo- the tree consists of the sapwood, which contains nesia, no equivalent detailed data are available. functional or living cells and the heartwood, which is formed by the death of cells. The species of fungi that are able to degrade the sapwood are generally dif- The heart-rot project ferent from those that can degrade the heartwood. The decay, which establishes itself in the heartwood Milestones and becomes progressive, is defined as heart rot. The project originated from within Australia but Heart rot has been reported to occur in the older has been based on the outcomes of an industry/ trees of many species of acacias, including Acacia research provider workshop held in Indonesia (Sub- auriculiformis A. Cunn. ex Benth. and A. mangium, anjeriji in Sumatra) in April 1995 that was partly sup- but its widespread occurrence in relatively young ported by ACIAR and CIFOR. trees of the latter species was unexpected. Heart rot The major research on heart rot of tropical acacias has not been reported from Acacia aulacocarpa has been carried out by Dr Lee Su See and her col- A. Cunn. ex Benth. or Acacia crassicarpa A. Cunn. leagues of the Forest Research Institute of Malaysia ex Benth. However, this may be because these two (Kuala Lumpur) and by Dr Shin-ichiro Ito of the species have not been systematically surveyed for Forest and Forest Products Research Institute (Kyoto, incidence of decay. Ito and Nanis (1994) found that Japan). There is clear evidence that branch stubs and A. mangium was more susceptible to heart rot than damage from singling are the main entry points of rot. was A. auriculiformis and a hybrid between these Dr Caroline Mohammed and co-workers at the Uni- species. versity of Tasmania have carried out the main body of Wounds are usually the entry points for the wood- work in Australia on entry of decay fungi via pruning decay fungi. On A. mangium trees, wounds include wounds. This has included pruning trials and research broken branches, mechanical injuries and branch on defence processes by Dr Karen Barry (University stubs after self-pruning or through singling and of Tasmania). Drs Inez Tommerup, Neale Bougher manual pruning operations. The incidence and extent and Morag Glen (CSIRO Forestry and Forest Prod- of the decay is highly variable and depends on many ucts, Perth) have carried out investigations in myco- factors including species, location of wound on the logical taxonomy and molecular studies.

23 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Objectives 1. Incidence of heart rots and root-rots in A. mangium (see Barry et al. 2004) Factors determining the extent of decay in acacias Surveys were completed at four sites (Perum Per- in Southeast Asia and eucalypts in Australian hard- hutani Jasinga, West Java; PT Wira Karya Sakti, wood have a high degree of commonality. This Jambi; PT Arara Abadi, Riau, East Kalimantan; and project sought to establish the means to minimise PT Musi Hutan Persada, South Sumatra), in which impacts of decay on the potential of A. mangium to trees were assessed at the harvest age of 8 years (in produce high-value products. The main aims of the addition, 3-year-old thinned trees were examined in project activities were to: Jasinga). A rapid method of surveying logs stacked in • assess the incidence of heart rot (and root rot) in the plantation was developed. different Indonesian environments The incidence of heart rot in the main stem was sig- • establish the relationship between silvicultural nificantly different between some regions, ranging practices and pruning of A. mangium with the from 6.7% in East Kalimantan up to 46.7% in West incidence of heart rot Java (Figure 1). However, the proportion of each • establish the relationships between trees selected defect type (1–4) did not show a consistent trend for other traits such as leaf pathogen/insect across all sites. A combination of differences in plan- resistance and form, and the incidence or risk of tation management (e.g. pruned or not pruned), age heart rot and climate between these five regions may explain • develop DNA-based methods coupled with the differences in heart-rot incidence and severity. traditional taxonomical and pathological methods to identify and characterise fungi causing rot 2. Effect of pruning on heart rot in A. mangium • assist plantation managers in Indonesia to reduce (see Beadle and Barry 2004). heart rot (and root rot) This experiment in South Sumatra set out to test • provide revised selection criteria for tree two hypotheses. The first was that pruning would be improvement projects in Indonesia to reflect associated with an increase in the incidence of heart outcomes of the research rot. Eighteen months after pruning, the incidence of • achieve technology transfer by staff exchange and heart rot was negligible or absent. As all the pruning interaction. wounds were occluded, it was likely that heart rot would not arise from pruning in this plantation in Outputs the future. Surveys of heat-rot incidence in harvested logs of trees aged 6–8 years had shown that it Some of the major outputs of the project are sum- varied with region in Indonesia (Barry et al. 2004). marised here. Other papers present the project While in South Sumatra the average incidence was outputs in more detail. low (11.3%), it was not absent. It was therefore dif-

Figure 1. Percentage of logs in each defect class (1–4) for the five survey sites. The numbers correlate with increasing severity of heart rot.

24 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. ficult to explain why heart rot was completely second survey. This highlights the potential for veg- absent in the trees assessed from the pruning trial. etative spread of the fungus after initial introduction The results suggested that interventions with to the site. pruning and thinning that are essential for the management of plantations for solid wood should not References increase the incidence of heart rot in South Sumatra if managed carefully. Almonicar, R.S. 1992. Two types of root rot diseases The second hypothesis tested whether form- affecting Acacia mangium. Nitrogen Fixing Tree pruning was associated with a lower incidence of Research Reports, 10, 94–95. heart rot and better form than was lift-pruning. The Arentz, F. 1996. Root rot in plantations of Acacia first part of the hypothesis could not be tested. How- auriculiformis and A. mangium. Proceedings of the ever, there was strong evidence that form-pruning IUFRO symposium on impact of diseases and insect pests was associated with better form, particularly in rela- in tropical forests, 98–104. tion to kinks, though only in the first 3 m of the stem Barry, K.M. and Irianto, R.S.B. 2004. Root rot surveys of where form-pruning had been undertaken. The Acacia mangium in Indonesia. In: Final research report results also suggested that form-pruning would have ACIAR project FST/00/123 Heartrots in plantation hardwoods in Indonesia and south-east Australia. been effective in the next 3 m of the stem had it been Barry, K.M., Irianto, R.S.B., Santoso, E., Turjaman, M., undertaken after the end of the third growing season Widyati, E., Sitepu, I. and Mohammed, C.L. 2004. (March–September 2003) when the trees were 8 m Incidence of heart-rot in harvest-age Acacia mangium in high (tree height was around 12 m at harvest). Indonesia, using a rapid survey method. Forest Ecology and Management, 190, 273–280. 3. Surveys for root rot of A. mangium in Indonesia Beadle, C. and Barry, K.M. 2004. Effect of pruning Acacia (Barry and Irianto 2004) mangium on heartrot, growth and form. In: Final research Of the diseases of A. mangium identified, root rot is report ACIAR project FST/00/123 Heartrots in plantation the most significant in terms of loss of productivity, hardwoods in Indonesia and south-east Australia. resulting in tree death (Old et al. 2000). Root rot of Ito, S.I. and Nanis, L.H. 1994. Heart-rot in Acacia mangium A. mangium plantations caused by Ganoderma spp. in Safoda plantations. SAFODA, JICA Sabah re- or Phellinus spp. has been found to occur throughout afforestation technical development and training project, Asia, with reports from Malaysia (Lee 1997), the 52p. Philippines (Almonicar 1992), Papua New Guinea Lee, S.S. 1997. Diseases of some tropical plantation acacias (Arentz 1996), and India (Prasad and Naik 2002). In in Peninsular Malaysia. In: Old, K.M., Lee, S.S. and Indonesia, the disease is also widespread (Rahayu Sharma, J.K., ed., Diseases of tropical acacias. 1999; Old et al. 2000), but there have been limited Proceedings of an international workshop, Subanjeriji, reports to date on its incidence. South Sumatra, 28 April–3 May 1996. Bogor, Indonesia, CIFOR Special Publication, 53–56. This study surveyed root-rot incidence and spatial Old, K., Lee, S.S., Sharma, J.K and Yuan, Z.Q. 2000. A arrangement in commercial plantations and trials. In manual of diseases of tropical acacias in Australia, SE second-rotation commercial plantations in Sumatra, Asia and India. Bogor, Indonesia, Center for International root-rot incidence recorded was 3–26%. The com- Forestry Research, 104p. partments surveyed in Riau province had signifi- Prasad, M. and Naik, S.T. 2002. Management of root rot and cantly higher root rot than those in South Sumatra. heart-rot of Acacia mangium Willd. Karnataka Journal of Fruitbodies of Ganoderma philippii and other Agricultural Science, 15, 321–326. species were collected in both regions during inde- Rahayu, S. 1999. Penyakit tanaman utan di Indonesia. pendent collections. In a provenance/family trial of Gejala, penyebab dan teknik pengendaliannya. A. mangium in Java, root-rot incidence was surveyed Yogyakarta, Penerbit Kanisius, 112p. [In Indonesian] at two separate times. Root rot incidence was about Zakaria, I., Wan Razali, W.M., Hashim, M.N. and Lee, S.S. 7% at the time of the first survey and 13% during the 1994. The incidence of heart-rot in Acacia mangium second survey, 1 year later. Spatial analysis by dis- plantations in Peninsular Malaysia. Kuala Lumpur, tance indices (SADIE) revealed that infected trees Forest Research Institute Malaysia, Research Pamphlet, were randomly distributed at the first time point but 114, 15p. tended to become aggregated by the time of the

25 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Heart rot and root rot in Acacia mangium: identification and assessment

Caroline L. Mohammed1, Karen M. Barry2 and Ragil S.B. Irianto3

Abstract

Heart rot in Acacia mangium is a typical white rot caused by hymenomycetes, which attack cellulose and lignin. Its development is associated with changes in colour, texture and appearance of rotted wood. These were used as the basis for the rapid assessment of the incidence and severity of heart rot on harvested log- ends in the field. The results are compared with previous survey reports based on longitudinal sectioning of logs. An assessment of two trials, one in Riau, the other in South Sumatra, showed that provenance could influence heart-rot incidence but that this was not correlated with wood extractive content. Root rots are characterised according to the colour of the infected fungal tissues/roots and are associated with various basidiomycetes. Molecular technology has identified Ganoderma philippii as the causative agent of red root rot. Root-rot diseases are associated with crown dieback, reduced growth and tree death. When root rot is first detected, infected trees or disease foci tend to be randomly distributed but then enlarge and may aggregate. The rate of disease progress appears positively related to the initial incidence of root rot. Accurate surveys to investigate spread should record above-ground symptoms, inspect the extent of root infection, and observe patterns of disease infection. However, these types of surveys, especially over extensive and difficult terrain, are operationally laborious and costly. Future options for disease survey based on remote sensing are discussed.

Surveys to evaluate diseases of tropical Acacia mangium Willd. is a saprotrophic fungal decay of plantations have concluded that heart rot, root rot heartwood which reduces wood quality but the tree and phyllode rust (respectively, infection of heart- is not killed and is, in most cases, externally wood, roots and leaves by fungi) are the main asymptomatic. Heart-rot fungi are wound basidio- threats (Old et al. 2000). Heart rot in Acacia mycete parasites that enter trees through injuries and branch stubs and do not preferentially attack living tissue. Root-rot disease in A. mangium is a 1 Ensis – the joint forces of CSIRO and Scion, Private Bag decay of roots caused by various basidiomycete 12, Hobart, Tasmania 7001, Australia; and University of pathogens, which attack living root tissue and may Tasmania, School of Agricultural Science, Private Bag 54, Hobart, Tasmania 7001, Australia. result in tree death or symptoms of crown decline. Email: . The disease is spread by the contact of a diseased 2 University of Tasmania, School of Agricultural Science, root or infested woody debris with a healthy root. Private Bag 54, Hobart, Tasmania 7001, Australia. Assessments of heart-rot and root-rot diseases Email: . require different approaches. For practical manage- 3 Ministry of Forestry FORDA, Centre for Forest and Nature Conservation Research and Development, Jalan ment in plantation forests, methods need to be fast, Gunung Batu No. 5, Bogor 16610, Indonesia. accurate and easily repeatable by staff with a Email: . minimum of training.

26 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Heart rot methods. In five major studies, the merchantable stem length was sectioned into 1 m logs that were each Internal symptoms of heart rot in sliced longitudinally to allow detection and quantifica- A. mangium tion of heart-rot incidence and severity (Mahmud et al. When healthy A. mangium trees are dissected, 1993; Zakaria et al. 1994; Ito and Nanis 1994; Basak sound sapwood is tan to bright yellow in colour, while 1997; Ito 2002). While destructive methods are sound heartwood is pale brown to grey brown. In extremely valuable for gaining exact measures of heart A. mangium the juvenile wood is pale coloured and of rot and understanding paths of fungal entry into the low density. It is located in the centre of the log, is stem (Lee et al. 1988; Mahmud et al. 1993; Ito and often soft, and can be mistaken for heart-rotted wood. Nanis 1994, 1997), they are labour intensive. This Like most fast-growing trees, A. mangium possesses a necessitates that replicate tree numbers within treat- high proportion of juvenile wood. It usually forms in ments are reasonably low; for example, 5–6 trees per the stem of the live crown and has been known to con- 13 plantations in Bangladesh (Basak 1997) or 10 trees stitute up to about 10% by volume of the wood present per 20 plots of different age in Sabah (Mahmud et al. in 8–9-year-old trees (Ivory 1993). 1993). For regular monitoring, a quicker method of heart-rot survey is desirable. The first symptom indicative of fungal infection is discoloration; heartwood becomes purple/black, Barry et al. (2004) developed a quick and easily sapwood becomes green/brown. Incipient decay in repeatable method of heart-rot survey to screen large heartwood is difficult to detect but the wood colour is numbers of logs in different regions of Indonesia. At intermediate between that of sound and decayed wood, most sites, logs were surveyed during harvest opera- i.e. it is usually of darker colour than normal heart- tions at the end of the rotation, generally at age 8 wood. Heart rot in A. mangium is typical of white rot years, but at age 6 years in East Kalimantan. In West caused by hymenomycetes (a group of basidiomycetes; Java, some trees were surveyed at harvest at age 8 Hood (2006)), which attack both cellulose and lignin, years (60 trees), and others at age 3 years during thin- finally leaving behind a yellowish-white, spongy or ning (39 trees). The number of compartments and stringy mass. Lee and Maziah (1993) described seven logs per compartment surveyed were based on avail- types of heart rot from A. mangium based on differ- ability of harvested material. ences in colour, texture and general appearance of the Log-ends were assessed where logs were stacked rotted wood. The authors attributed each type to dif- after harvest. Typically, approximately 100 logs ferent stages of decay as well as characteristics of the were in each pile and 3–4 logs could have been individual fungi involved. Isolated decay-columns may derived from an individual tree. For example, in East form in the heartwood. In extreme cases, heartwood Kalimantan, logs were about 3.5 m long and 3–4 logs can completely rot away leaving a hollow core. of this length were produced from an individual tree. The infection courts for heart-rot fungi in Therefore, logs in the piles were not uniform in size A. mangium are dead branch stubs, dead/broken and could have come from a number of positions in branches and stem cankers/unhealed or slowly healing the tree. The logs chosen to assess from each log pile wounds such as those caused by pruning (Mahmud et were selected randomly by using a line transect (a al. 1993; Ito and Nanis 1994, 1997; Barry et al. 2004). piece of string placed horizontally at random), which Infection courts (i.e. branch stubs) are not often associ- typically selected 10–15 logs (Figure 1). Since A. ated with external signs or symptoms such as fruit- mangium wood discolours rapidly upon cutting, logs bodies, and heart rot can be seen only after the bolts of were assessed as soon as possible after harvest. Dis- wood are split. Molecular techniques have greatly coloration related to heart rot can appear similar to increased the ability to identify isolates obtained from oxidative discoloration, and this is therefore a poten- diseased wood by matching the DNA of such isolates tial source of confusion. The diameter of each log to that of named fruitbodies (Glen et al. 2004). selected was measured and if a heart-rot-related defect was observed this was categorised on a 1–4 Methods of detection and assessment scale (Figure 1) and its maximum diameter measured. Logs with no heart rot were scored as 0. Where Since it is not possible to assess heart rot from the more than one defect type was present on one log- external features of the living tree, most surveys of end, the highest rating was recorded and the widest A. mangium heart rot have utilised destructive diameter of the total defect was recorded.

27 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9February 2006. Canberra, ACIAR Proceedings No. 124.

Figure 1. Heart-rot defect scale and log stack showing line transect method of Barry et al. (2004)

Comparing results of the log-stack method to of heart rot than those found by Barry et al. (2004) in other studies Indonesia. This difference could be due to the method used in Indonesia, which may underestimate heart- In the Indonesian study by Barry et al. (2004), the rot incidence as it examines only one cross-section of East Kalimantan and South Sumatra sites were asso- each log (although it surveys a larger number of ciated with the lowest heart-rot incidence (Figure 2) logs). As such, it is not possible to compare the and the lowest average defect diameters. Lower results from Barry et al. (2004) with surveys con- heart-rot incidence would be expected for the East ducted by other methods. Surveys by Maurits et al. Kalimantan site, mainly because the trees were 2 (2001) found volume losses to heart rot of 0.7–14.1% years younger than at the other sites (Figure 2). This in A. mangium in East Kalimantan. Correlating the age-related trend has been evident in other studies of different methods would require a destructive anal- A. mangium, with positive correlations between ysis of a sub-sample of logs following the survey of heart-rot incidence and tree age (Zakaria et al. 1994) the same logs with Barry et al.’s method. However, it as well as between heart-rot diameter and tree diam- is very possible that there are real differences in inci- eter (Basak 1997). Apart from tree age, other factors dence between the various studies. For example, seed such as climate and management practices may influ- stock has improved markedly in the 15 years between ence heart-rot incidence and spread. the first Malaysian studies and the more recent Indo- nesian surveys (C. Harwood, pers. comm.) and these improvements may also reflect an associated reduction in susceptibility to heart rot; trees have become straighter, branches smaller and less numerous, thus reducing the frequency of suitable infection courts presented by broken or dead branches, especially those of large diameter (Lee et al. 1988; Mahmud et al. 1993; Ito and Nanis 1994, 1997; Lee 2002).

Assessment of intra and inter specific susceptibility to heart rot in acacia Figure 2. Percentage of logs in each defect class It has been suggested that provenance affects sus- (1–4) for the five survey sites ceptibility to heart rot (Ito and Nanis 1994). As mentioned above, provenance differences in tree Previous Malaysian surveys using destructive architecture, especially branching, may influence methods of analysis have detected higher incidences heart rot. A comparison of polyphenols between

28 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

A. mangium (heart-rot susceptible) and A. auriculi- noxius (brown rot), Rigidoporus lignosus (white rot) formis (heart-rot resistant) showed that some compo- and Tinctoporellus epimiltinus (brown rot) (Almon- nents are toxic to heart-rot fungi when at certain icar 1992; Old et al. 2000; Lee 2002; Glen et al. concentrations (Barry et al. 2005; Mihara et al. 2005). 2006). Species of Ganoderma, however, have been Wood extractive content may also vary intra-specifi- reported as the most frequent root-rot causal agents in cally with provenance and result in differing levels of A. mangium in both Malaysia and Indonesia (Lee heart rot. 2002). Until now, identification of the Ganoderma Two trials clearly demonstrated that the effect of species associated with root disease has been based provenance was statistically significant for heart-rot on morphological and taxonomic features of fruit- incidence (Barry et al. 2006). The larger study in Riau bodies (Figure 3a). Roots affected by Ganoderma assessed natural heart-rot incidence by felling trees spp. may be covered by a reddish-brown rhizomor- and assessing a stem disk at 1.5 m height. In the phic skin (Figure 3b) that is visible after the roots are smaller provenance trial in South Sumatra, trees were washed clean of associated soil. Using molecular artificially inoculated. This involved creating stem technology, Ganoderma philippii was unequivocally wounds (drill holes), which were then inoculated with identified as a causative agent of red root rot (Glen et fungi capable of causing heart rot. This method al. 2004, 2006). A white mottling pattern of myc- offered a controlled way of testing for provenance elium is seen on the underside of the infected bark susceptibility, as the fungal species inoculated and the (Figure 3b) and there is a very characteristic odour. In size of the infection court were standardised. How- fast-grown plantations, characteristic fruitbodies ever, a range of other fungi infected the wounds after may be absent in disease centres (Lee 2000; Old et al. inoculation. While both trials revealed that prove- 2000). A species of the G. lucidum complex is asso- nance could influence heart-rot incidence, wood ciated with a more rubbery rhizomorphic skin than extractive content was not correlated with heart rot. that found in red root rot, a skin which is yellow and brown on the outside with a blistered appearance and Root rot rubbery white on the inside when pulled away from the root wood (M. Glen, pers. comm.). Signs Symptoms Several types of woody root rots have been recognised in A. mangium and are characterised as red, Most root-rot diseases cause similar, generally non- brown, black or white root diseases according to the specific, above-ground symptoms in the host, colour of the fungal tissue and/or roots that are including crown dieback and reduced growth. There infected. These diseases are associated with various is a general decline in the crown condition and the basidiomycete fungi including Amauroderma cf. growth rate is poor. In particular, the foliage of parasiticum, Ganoderma spp. (red rot), Phellinus affected trees becomes chlorotic (paling of the green

(b) (a)

Figure 3. (a) Ganoderma philippii basidiocarp at the base of a dead Acacia mangium tree growing in East Kalimantan, Indonesia; (b) characteristics of red root rot (red bumpy skin when soil is washed away) caused by G. philippii on A. mangium with white mottling of mycelium seen on underside of bark

29 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9February 2006. Canberra, ACIAR Proceedings No. 124. coloration), much reduced in size and sparse due to plantation. It is therefore useful to investigate effi- reduced water and mineral uptake. Young shoots may ciencies of different survey methods to save moni- wilt and, in trees in advanced states of root rot, foliage toring time and costs. loss increases and trees become very susceptible to Some surveys have utilised a mixture of above- wind throw (Ariffin et al. 2000; Kile 2000; Leckie et ground symptoms and root-infection inspections. al. 2004; Morrison et al. 2000; Old et al. 2000). Irianto et al. (2006) conducted a survey at different In forest stands, diseased trees tend to be clustered sites in Indonesia by recording the incidence of root in patches that are roughly circular, due to the spread rot in two adjacent rows of A. mangium, missing the of root rot from around an initial inoculum source next three rows and then repeating this survey pattern (Figure 4). Trees with advanced infections are until a certain count was reached to represent a 40% located at the infection centre, while trees with less- sub-sample of a randomly selected area of the com- advanced infections are around the periphery. These partment. In East Kalimantan, for example, trees circular disease centres increase in diameter as root were assessed until a count of 200 was reached, the contact occurs between neighbouring diseased and count therefore representing a 40% sub-sample of healthy trees (Bolland 1984; Garbelotto et al. 1997; 500 trees. Roots were uncovered and inspected for Kile 2000). typical symptoms of root rot (red) in trees that were standing dead, or exhibited chlorotic/yellowing Quantitative detection using ground-based foliage, loss of foliage, or fruitbodies on the stem. surveys Trees in which root rot was confirmed by this Surveys are essential as a management tool to method, plus any missing trees, were scored as a pos- determine if disease incidence is severe enough to itive result. Missing trees were attributed to root rot. warrant control practices, but also as a study tool, or Results were recorded as presence/absence of root for epidemiological modelling. Surveys can utilise rot, then the numbers in each of the two categories the above-ground symptoms of root rot (e.g. crown were summed and expressed as a percentage. During condition, resin exudates on bark) or involve field surveys, observations on the pattern of root-rot checking for signs of root infection by excavating infections (i.e. size of root-rot disease foci) were around roots. Field-based surveys of root rot can be recorded. While this survey method was time-con- completed quickly for small areas or trials, but are suming, it ensured a very accurate assessment of time-consuming at the large scale of a commercial root-rot incidence.

Figure 4. Typical circular patch of tree mortality (3–5 trees) in a 4–5-year- old Acacia mangium plantation, East Kalimantan, Indonesia

30 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Surveys are useful not only to provide quantitative 2004). While remote sensing methodologies require data from one time point but also as a basis for model considerable development for reliable detection of development (Nandris et al. 1996). Barry et al. (2005) specific diseases, they offer the potential for rapid modelled six years of spatial survey data of rubber trees and robust quantification. The symptoms of root rot infected with Rigidoporous lignosus and Phellinus in A. mangium are suitable for adaptation to remote noxius. Using the spatial information that was collected sensing. while testing different control methods, models were determined to estimate root-rot spread and effective- Conclusions ness of control measures. Successive surveys conducted in A. mangium plantations infected by For both of these important diseases of A. mangium, Ganoderma spp. (Lee 2000) showed that the rate of assessment methods need to be rapid and accurate. disease progression in Malaysian plantations was Heart rot requires destructive assessment, but this can dependent on the initial incidence of root rot. Root-rot be done during thinning operations or at harvest. Root foci enlarged over time. rot can be assessed non-destructively but root excava- Assessment of root rot in an A. mangium prove- tion is required for verification of the disease. Root- nance/progeny trial in Indonesia (Irianto et al. 2006) rot assessment methodology is therefore both labo- revealed that infected trees were randomly distributed rious and costly, more suited to research investiga- at a first time point but these trees or disease foci tended tions which do not necessarily require full spatial to become aggregated by the time of a second survey. coverage of a forest plantation estate. Remote sensing This highlighted the high probability that exists of rapid of root-rot incidence and severity could offer 100% vegetative spread by the fungus after its initial intro- coverage. Knowledge of disease incidence and duction to the site; the incidence of tree death doubled severity (and patterns of spread in the case for root rot) between 2003 and 2004. While this provenance/ is important for developing effective disease manage- progeny trial is a non-commercial plantation, the ment strategies and for calculating economic impact. results have implications for the potential rate of spread of root rot in plantations. Lee (2000) found that for first- References rotation plantations grown on lowland ex-forest sites, mortality due to root rot in A. mangium in Malaysia Almonicar, R.S. 1992. Two types of root rot disease could double within one year. The incidence of other affecting Acacia mangium. Nitrogen Fixing Tree root-rot fungi such as Armillaria ostoyae in pine forests Research Reports, 10, 94–95. has also been reported to double within one year (Lung- Ariffin, D., Idris, A.S., and Singh, G. 2000. Status of Escarmant and Guyon 2004). Ganoderma in oil palm. In: Flood, J., Bridge, P.D. and Holderness, M., ed., Ganoderma diseases of perennial Detection methods for the future crops. Wallingford, Oxon, UK, CAB International, 49– 69. As ground surveys are time-consuming, aerial or Barry, K.M., Irianto, R.S.B., Santoso, E., Turjaman, M., remote methods may be suitable for detection of root Widyati, E., Sitepu, I. and Mohammed, C.L. 2004. rot. Remote sensing of plant health is currently based Incidence of heartrot in harvest-age Acacia mangium in mainly on detection of chlorophyll content. Foliage Indonesia, using a rapid survey method. Forest Ecology thinning and chlorosis are common symptoms of root and Management, 190, 273–280. rot and, in Douglas-fir, chlorophyll a, nitrogen and Barry, K.M., Irianto, R.S.B., Tjahjono, B., Tarigan, M., moisture were consistently reduced in trees infected Agustini, L., Hardiyanto, E.B. and Mohammed, C.L. 2006. Variation of heartrot and extractives with by Phellinus weirii (Thomson et al. 1996). In beech provenance of Acacia mangium. Forest Pathology, 36, trees infected with the root rot caused by Phytoph- 183–197. thora spp., total chlorophyll was reduced three-fold Barry, K.M., Mihara, R., Davies, N.W., Mitsunaga, T. and in heavily infected plants compared with controls Mohammed, C.L. 2005. Polyphenols in Acacia mangium (Fleischmann et al. 2004). The application of a high- and A. auriculiformis heartwood with reference to resolution multi-spectral imagery was recently heartrot. Journal of Wood Science, 51, 615–621. shown to be a feasible for detecting laminated root- Basak, A.C. 1997. Heart rot of Acacia mangium in rot centres in Douglas-fir in Canada (Leckie et al. Bangladesh. Indian Journal of Forestry, 20, 61–66.

31 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9February 2006. Canberra, ACIAR Proceedings No. 124.

Bolland, L. 1984. Phellinus noxius: cause of a significant Leckie, D.G., Jay, C., Gougeon, F.A., Sturrock, R.N. and root-rot in Queensland hoop pine plantations. Australian Paradine, D. 2004. Detection and assessment of trees with Forestry, 47, 2–10. Phellinus weirii (laminated root rot) using high resolution Fleischmann, F., Gottlein, A., Rodenkirchen, H., Lutz, C. multi-spectral imagery. International Journal of Remote and Osswald, W. 2004. Biomass, nutrient and pigment Sensing, 25, 793–818. content of beech (Fagus sylvatica) saplings infected with Lee, S.S. 2000. The current status of root diseases of Acacia Phytophthora citricola, P. cambivora, P. pseudosyringae mangium Willd. In: Flood, J., Bridge, P.D. and and P. undulata. Forest Pathology, 34(2), 79–92. Holderness, M., ed., Ganoderma diseases of perennial Garbelotto, M., Slaughter, G., Popenuck, T., Cobb, F.W. crops. Wallingford, Oxon, UK, CAB International, 71– and Bruns, T.D. 1997. Secondary spread of 79. Heterobasidion annosum in white fir root-disease — 2002. Overview of the heartrot problem in acacia – gap centers. Canadian Journal of Forest Research, 27, 766– analysis and research opportunities. In: Barry, K., ed., 773. Heartrots in plantation hardwoods in Indonesia and Glen, M., Tommerup, I., Abou Arra, S., Bougher, N., Lee, Australia. Canberra, ACIAR Technical Report 51e, 18– S.S., Barry, K.M., Irianto, R.S.B. and Mohammed, C.L. 21. 2004. Identification of polypore fungi causing root rot Lee, S.S. and Maziah, Z. 1993. Fungi associated with heart and heart rot of Acacia mangium. 4th Asian Mycological rot of Acacia mangium in Peninsular Malaysia. Journal of Congress, Chiang Mai, Thailand, 14–19 November 2004. Tropical Forest Science, 5, 479–484. Glen, M., Bougher N., Nigg S.Q., Lee S.S., Irianto, R. and Lee, S.S., Teng, S.Y., Lim, M.T. and Razali Abdul Kader. Mohammed, C.L. 2006. Molecular differentiation of 1988. Discoloration and heartrot of Acacia mangium Ganoderma and Amauroderma species and their role in Willd. — some preliminary results. Journal of Tropical root disease of Acacia mangium plantations in Indonesia Forest Science, 1, 170–177. and Malaysia. Australasian Plant Pathology, in press. Lung-Escarmant, B. and Guyon, D. 2004. Temporal and Hood, I.A. 2006. The mycology of the Basidiomycetes. In: spatial dynamics of primary and secondary infection by Potter, K., Rimbawanto, A. and Beadle, C., ed., Heart rot Armillaria ostoyae in a Pinus pinaster plantation. and root rot in tropical Acacia plantations. Canberra, Phytopathology, 94, 125–131. ACIAR Proceedings No. 124, 57–68. [These proceedings] Mahmud, S., Lee, S.S. and Ahmad, H.H. 1993. A survey of Hood, I.A., Redfern, D.B. and Kile, G.A. 1991. Armillaria in heartrot in some plantations of Acacia mangium Willd. in planted hosts. In: Shaw, C.G. III, and Kile, G.A., ed., Sabah. Journal of Tropical Forest Science, 6, 37–47. Armillaria root disease. Washington DC, United States Maurits, S.S., Suyadi, Wahyu Wardana, Sugeng Widodo Department of Agriculture, Forest Service, Agricultural and Solikin, M.N. 2001. Incidence level of Acacia Handbook, No. 691, 122–150. mangium heart rot on forest plantation in Sumalindo Irianto, R.S.B., Barry, K.M., Hidayah, I, Ito, S., Fiani, A., Group East Kalimantan. Research Report, Research and Rimbawanto, A. and Mohammed, C.L. 2006. Incidence Development Planting Division, Sumalindo Group, and spatial analysis of root rot of Acacia mangium in Samarinda, 70p. Indonesia. Journal of Tropical Forest Science, 18, 87–95. Mihara, R., Barry, K.M., Mohammed, C.L. and Mitsunaga, Ito, S. 2002. The infection of heartrot and disease severity on T. 2005. Comparison of antifungal and antioxidant several Acacia species in SAFODA plantations. Study activities of Acacia mangium and A. auriculiformis report of SAFODA–JICA project, 16p. heartwood extracts. Journal of Chemical Ecology, 31, Ito, S. and Nanis, L.H. 1994. Heartrot of Acacia mangium in 789–804. SAFODA plantations. Sabah Reafforestation Technical Morrison, D.J., Pellow, K.W., Norris, D.J. and Nemec, Development and Training Project, study report. Sabah, A.F.L. 2000. Visible versus actual incidence of Malaysia, Chin Chi Printing Works, 52p. Armillaria root disease in juvenile coniferous stands in — 1997. Survey of heartrot on Acacia mangium in Sabah, the southern interior of British Columbia. Canadian Malaysia. Japan Agricultural Research Quarterly, 31, 65– Journal of Forest Research, 30, 405–414. 71. Nandris, D., Chadoeuf, J., Pierrat, J.C., Joannes, H., Geiger, Ivory, M.H. 1993. Status of forest plantations and potential J.P. and Nicole, M. 1996. Modelling rubber-tree root wood availability. In: Proceedings of the seminar on the diseases, simulations of various inoculum rates and potential of acacia and other plantation species, 15 methods of control. European Journal of Forest December 1993, Kuala Lumpur, 21–29. Pathology, 26, 25–44. Kile, G.A. 2000. Woody root rot of eucalypts. In: Keane, Old, K.M., Lee, S.S., Sharma, J.K. and Yuan, Z.Q. 2000. A P.J., Kile, G.A., Poupard, P. and Brown, B.N., ed., manual of diseases of tropical Acacias in Australia, Diseases and pathogens of eucaypts. Collingwood, South-East Asia and India. Jakarta, Indonesia, Centre for Australia, CSIRO, 293–306. International Forestry Research.

32 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Thomson, A.J., Goodenough, D.G., Barclay, H.J., Lee, Y.J. Zakaria, I., Wan Razali, W.M., Hashim, M.N. and Lee, S.S. and Sturrock, R.N. 1996. Effects of laminated root rot 1994. The incidence of heartrot in Acacia mangium (Phellinus weirii) on Douglas-fir foliar chemistry. plantations in Peninsula Malaysia. Forest Research Canadian Journal of Forest Research, 26, 1440–1445. Institute Malaysia, Research Pamphlet, 114, 15p.

33 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

The mycology of the Basidiomycetes

Ian A. Hood1

Abstract

A number of significant tree and plantation root diseases in tropical Asia are caused by certain species of basidiomycetes. This important group of fungi, which includes such familiar forms as mushrooms, toadstools, bracket, shelf, and crust fungi, puff balls, earthstars, and coral fungi, is characterised by the production of sexual spores on the outside of a microscopic structure called the basidium. Basidiomycetes occupy many niches in the environment, including decomposing litter, decaying wood and soil organic matter, in a variety of habitats such as forests and open countryside. Some species form beneficial mycorrhizal relationships with the roots of host trees, but others are important pathogens of the foliage, stems or root systems of different tree species. Basidiomycete species are traditionally identified by the form and microscopic structure of their fruitbodies, and by their appearance when isolated in laboratory culture. Important root diseases of trees in Indonesia are caused by the basidiomycete fungi Rigidoporus microporus, Junghuhnia vincta, Phellinus noxius, and certain species of Ganoderma.

Mycology is the scientific study of fungi. It is distin- (slime moulds) and the oomycetes (which feature the guished from, but related to plant pathology, the downy mildews and microscopic root-infecting fungi study of plant diseases, a large proportion of which such as species of Phytophthora and Pythium). Most are caused by harmful fungi (fungal pathogens). of the true fungi exist and grow in the form of a Fungi are a very large, diverse group of living spreading network or mycelium of living tube-like organisms found in nearly all ecosystems. Fungi are threads called hyphae (though yeasts develop as eukaryotes, that is, they have microscopic organelles budding cells). The hyphae branch as they grow from within their cells called nuclei which contain genetic the tip, and feed heterotrophically. This means that material in the form of thread-like chromosomes , fungi cannot photosynthesise like plants and algae, enabling hereditary characters to be passed on to sub- which are autotrophic, but live either saprobically on sequent generations. Other eukaryotes include all dead organic matter or symbiotically in association members of the plant and animal kingdoms, whereas with other living organisms. The latter relationship life forms such as bacteria and blue–green algae, (symbiosis) may be harmful to the living plant or whose genetic material is not held in a nucleus, are animal host (parasitic), mutually beneficial (as in called prokaryotes (Table 1). lichens and mycorrhizas), or neutral (commensal, as Recent study has shown that some of the organ- in fungi that grow as endophytes within living plant isms we call fungi actually belong to other groups. leaf, root or stem tissues without causing harm). Those no longer recognised as true fungi, include Fungi reproduce and spread by means of sexual fungus-like organisms such as the myxomycetes and asexual spores, both of which may be released in large numbers. In one of the two largest groups of 1 Ensis – the joint forces of CSIRO and Scion, Te Papa true fungi, the ascomycetes , sexual ascospores are Tipu Innovation Park, Private Bag 3020, Rotorua, New produced within a sac-like structure called the ascus. Zealand. Email: . Among the larger ascomycetes are the disc and cup

34 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. fungi and a specialised category of fungi called the taxonomy has been controversial because the number lichens that form a symbiotic relationship with of good distinguishing characters is limited, and there certain green algae. In the second large group, the is uncertainty and disagreement as to which features basidiomycetes, sexual basidiospores are produced are more important for separating the different sec- on the outside of a modified cell called the basidium. tions and even species. The recent upsurge in DNA Ascomycetes such as Kretzschmaria zonata (syn- technology has provided a powerful addition to tradi- onym, Ustulina zonata; sometimes incorrectly as tional taxonomic methods. U. deusta) and Corallomycetella repens (synonym, Most mycologists separate two major groups, the Sphaerostilbe repens) may attack roots of hardwood ‘rusts’ and the ‘smuts’, from among the remaining crops such as rubber, tea or cacao, but basidiomycete basidiomycetes (Table 2). Both contain a large fungi are the most common causes of root diseases number of specialised microscopic parasitic fungi and decay in natural forests and plantations. whose mycelium grows within the foliage, twig or stem tissues of flowering plants, conifers or ferns. The nature of the basidiomycetes They have complicated life cycles featuring, in the case of the rusts, up to five different spore types that The basidiomycetes make up a huge variety of fungi allow the fungus to alternate between different hosts. (ca. 23,000 species are known). Determining their However, many rust species have reduced life cycles

Table 1. How the basidiomycetes relate to other living organisms

– prokaryotes (nucleus absent) – bacteria – blue–green algae – other prokaryotes – eukaryotes (nucleus present) – animals – plants and red and green algae – slime moulds (several groups) – water moulds (oomycetes) and brown algae – true fungi – ascomycetes (including lichens) – basidiomycetes – other true fungi once grouped in the former ‘phycomycetes’ (glomeromycetes, zygomycetes, chytridiomycetes) – other eukaryotes

Table 2. A simplified, artificial classification of the basidiomycetes

– rusts – smuts – others – heterobasidiomycetes (divided basidium; e.g.wood ears, jelly fungi) – homobasidiomycetes (simple, undivided basidium) – ‘gasteromycetes’ (e.g. puff balls, earth stars, stink horns, birds-nest fungi) – ‘hymenomycetes’ – Dacrymycetales – ‘Agaricales’ (broad sense, e.g. mushrooms, toadstools, boletes) – ‘Aphyllophorales’ (e.g. bracket and shelf fungi, corticioid fungi, toothed and spined fungi, coral fungi) – Others

35 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. that lack some spore states and occur on only one host. variety of only distantly related families, they too are The remaining basidiomycetes, the ‘others’ (Table 2), still in use as convenient, all-embracing, informal include the larger forest fungi e.g. mushrooms, toad- categories. In the agarics, the basidia are produced on stools, jelly and bracket fungi. Many members of this the gill surfaces. The spores, when released, fall group are linked in common by a unique type of under gravity, emerge from between the vertically septum (the cross wall that divides the hyphae into orientated gills, and are blown away in the wind. The separate cells) known as the dolipore septum. These Aphyllophorales comprise many different forms fungi fall into two sub-groups; the heterobasidiomyc- including horizontal bracket and shelf fungi, sheet or etes, in which the basidium initial is partitioned by crust fungi (corticioid), toothed fungi (with vertically walls into four cells following meiosis (Figure 1), and hanging spines or teeth), and coral fungi (with erect the homobasidiomycetes, in which it remains undi- antler- or tree-like branches). The life cycle of a vided as a single cell. typical aphyllophoroid basidiomycete, showing also The homobasidiomycetes were once conveniently a typical simple, homobasidiomycete basidium, is separated into the hymenomycetes, whose spores are portrayed in Figure 2. A comparatively small number actively ejected from the basidium in the expanded of basidiomycete species also produce asexual spores fruitbody (basidiocarp), and the gasteromycetes, (e.g. the important Northern Hemisphere root disease whose spores are held passively. It is now generally fungus, Heterobasidion annosum). accepted that both groups are artificial, being composed of a heterogeneous collection of different fam- Basidiomycetes in the environment ilies, though for practical purposes they are still used in an unofficial way as helpful, readily understood Like other fungi, different basidiomycetes feed categories. The gasteromycetes include the puffballs saprobically or symbiotically. Saprobic basidiomyc- and earth stars, which rely on the impact of falling etes occur in forests (in the soil, in leaf or twig litter, rain drops to puff their spores into the air; the or as wood colonisers in standing trees or fallen unpleasantly smelling stinkhorn; veil and basket stems), in open country (also in soil or decomposing fungi that attract flies for spore dispersal; and the litter), and in many other places. Many basidio- birds nest fungi that also depend on the force of mycete fungi serve an important ecological role as falling rain to splash their ‘eggs’ (packets of spores) wood decomposers (see Mohammed et al. 2006). A over a considerable distance. The hymenomycetes number of small toadstool species occupy a special- incorporate the Dacrymycetales (small, brightly col- ised position, along with other fungi, in colonising oured, jelly-like fungi that can be rewetted and reju- animal dung (coprophily), and species of Termito- venated after drying down, and which have a forked, myces (a genus of toadstool fungi) are found within two-spored basidium), the traditional Agaricales termite nests. (also known as agarics — gilled mushrooms and Another important ecological niche is filled by toadstools, and poroid boletes), and the Aphyllopho- certain agaric species (mushrooms and toadstools) rales. Although both the Agaricales and Aphyllopho- and also other types of subterranean basidiomycete rales are also heterogeneous groups containing a (e.g. Rhizopogon species) which beneficially infect

Figure 1. (A) Fruitbody, (B) septate basidia, (C) detached spore, and (D) clamped hyphae of Auricularia spp. (wood ear fungi), within the heterobasidiomycetes. After Hood (2003)

36 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. the small roots of living trees forming an ectomycor- species belonging to Crinipellis and Marasmius are rhizal association (one of several types of mycor- associated with leaf and twig blights in tropical forest rhiza). In this relationship, the fungus benefits by canopies and understoreys. Crinipellis perniciosa receiving carbohydrates from the host, while the and Mycena citricola cause witches broom and leaf mycelial network extending out into the soil from the spot diseases of cacao and coffee, respectively, in infected root system provides the host with an central and South America. Rusts and smuts cause enhanced supply of nitrogen and phosphorus, partic- serious diseases of major food crops. Rusts infecting ularly advantageous in nutrient depleted soils. Acacia species include Racospermyces digitatus (synonym, Atelocauda digitata), which causes dis- tortion and malformation of the phyllodes of species such as A. mangium, and Uromycladium notabile and U. tepperianum, which produce globose galls and eventually dieback on bipinnate and phyllodinous acacias. In temperate zones, important root diseases of plantation forests are caused by species in the Het- erobasidion annosum complex and by species of Armillaria. Armillaria root disease often has more impact in cooler, higher elevation forests in the tropics, while at lower altitudes, significant root diseases are caused by Rigidoporus microporus (synonym, R. lignosus), Junghuhnia vincta (synonym, R. vinctus), Phellinus noxius, and species of Gano- derma.

Identifying basidiomycetes

The body or thallus of the basidiomycete fungus (the Figure 2. Lifecycle of a typical hymenomycete. mycelium) is normally hidden within the substrate, Plasmogamy is the fusion of one cell from and it is generally only the fruitbody or basidiocarp each of the paired primary mycelia that is visible at the surface. For this reason, and without union of their nuclei; as a because the fruitbody tends to show the greatest mor- consequence, the new cells in the resultant phological variation, conventional mycology relies secondary mycelium each contain two on a number of macroscopic and microscopic fea- haploid nuclei, n + n. Karyogamy is tures of the fruitbody to distinguish between species. nuclear union resulting in a single A selection of the more usual of these characters is combined diploid, 2n, nucleus within the cell; the haploid state is restored by catalogued below, concentrating particularly on the nuclear division through the process of Aphyllophorales (e.g. corticioid and polypore fungi), meiosis in the basidium, allowing re– which are more likely to be associated with root assortment of the genetic material among disease and wood decay in hardwood plantations. the progeny. After Gadgil (2005) The use of fruitbody morphology in distinguishing between fungi is sometimes augmented by other Certain parasitic basidiomycete fungi cause impor- methods such as the appearance and form of the tant tree diseases. Examples of stem disorders fungus in pure culture (see below); physiological fac- include, in the tropics, pink disease (caused by Eryth- tors, such as the degree of virulence to different hosts, ricium salmonicolor; synonyms, Corticium salmoni- environmental growth preferences (nutrients, tem- color, Phanerochaete salmonicolor) and in perature); and by relationships at the molecular level, temperate regions, silver leaf disease (caused by as shown in protein profiles (e.g. immunological or Chondrostereum purpureum). A number of agaric isozyme analysis) and DNA sequencing.

37 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Macroscopic features of the basidiocarps confirm the identification. Some of the microscopic characters include the following: Longevity: annual or perennial (surviving one or more than one year). Hyphal composition of fruitbody tissues (Figure 3): In simple terms there are three types of hyphae Texture: soft and fleshy, gelatinous, cartilaginous, present in most corticioid and polypore fungi. brittle, corky, leathery, or woody; dry, moist, or sticky; upper surface smooth, velvety, hairy or scaly. Generative hyphae: thin, or occasionally thick- walled, branched hyphae, with septa (cross walls) Colour of internal tissues (context and spore- found in all fruitbodies. Called generative, because bearing component): white, pale brown, dark brown, they give rise in some species to other types of or other (e.g. black, red). hyphae. Septa may be simple (i.e. lack clamps) or Form: pileate (projecting out from the substrate with one, or sometimes more structures called clamp surface), resupinate (forming a flat sheet or crust), or connections at each cross wall (the clamp connection effused-reflexed (forming a shelf with the base is a hyphal outgrowth that bridges two adjacent cells extending down over the substrate as a flat sheet); if resulting from cell division, enabling one of the pileate: simple or compound; stalked (stipitate) or daughter nuclei to pass from one cell to the other; sessile; solitary or clustered (possibly imbricate, i.e. clamps are unique to, and help to identify many several shelves overlapping one above the other); if basidiomycetes, but not all basidiomycetes have stalked, attached at the side or centrally. clamps; illustrated in Figures 1, 2 and 3). Spore and basidia-bearing surface: smooth, Skeletal hyphae: long, thick-walled or solid, folded or warty, usually forming the fruit body under unbranched (or sparingly branched in some species) surface (e.g. corticioid fungi); lining vertical gills or hyphae lacking septa, derived from generative lamellae (e.g. agaric fungi; Lenzites , Panus, Pleu- hyphae. rotus species); lining vertical, downward-directed Binding hyphae: much-branched, thick-walled or pores (e.g. polypore fungi; boletes); on vertically solid hyphae lacking septa, derived from generative hanging teeth or spines (hydnoid fungi); on erect hyphae. May be narrower than skeletal hyphae. branches (e.g. ramarioid fungi) etc. These hyphal types allow for three kinds of fruit- Dimensions: size of fruitbody; width of context, body composition, which are characteristic of partic- pores, lamellae. ular genera and species: Host: the identity or type of host supporting a fruc- Monomitic: composed only of generative hyphae tification (fruitbody) may be important. (which in monomitic construction also provide sup- Nature of decay associated with a basidiocarp on port; characteristic, therefore of many non-woody wood: white, fibrous or stringy (simultaneous delig- fruitbodies). nification); brown, cubically fractured; pocket rot Dimitic: composed of generative and either skel- (selective delignification); appearance of any zone etal or binding hyphae (which provide the support). lines (pseudosclerotial plates). Trimitic: composed of all three types of hyphae The fruitbodies of agaric basidiomycetes (mush- (support being provided by skeletal hyphae bound rooms and toadstools) are distinguished initially by together by interweaving binding hyphae). their spore colour (as revealed in the spore deposit); Dimitic and trimitic types of construction are char- the presence or absence of a ring around the stalk or acteristic of more woody, perennial fruitbodies, in stipe, and a cup-shaped structure called the volva at which generative hyphae are more easily found at the its base (the partial and universal veils); the nature of still developing areas such as the fruitbody margin, the substrate (soil or wood); surface texture (smooth, hymenium (spore-producing layer), or pore mouth. velvety, scaly; dry, sticky, with fibrils etc.); and the When identifying species, it is also important to form of the gill attachment to the stipe. record the dimensions, colour, and colour reactions of the different types of hyphae. Microscopic features of the basidiocarps Nature of the hymenium: The macroscopic form of basidiomycete fruit- The hymenium is the microscopic layer containing bodies may vary considerably according to situation, the basidia and spores. Species can be distinguished and the overall appearance can sometimes be mis- by the form and size of the basidium (including the leading without a microscopic examination to number of sterigmata or appendages that bear the

38 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. spores, usually four, and whether there is a clamp Identification in culture present at the septum at the base), the presence and character of other structures, such as cystidia, metu- Very few basidiomycete fungi fruit in artificial loids (lamprocystidia) and cystidioles (Figure 3), and culture, so although different species can be distin- the appearance of the basidiospores, if present. guished from one another by recording their vegeta- Spores are distinguished by their shape and size, tive appearance and structure during growth, it is surface ornamentation (smooth, warted etc.), colour, frequently not possible to name them, even when and colour reaction to an iodine-based chemical comparison is made with a number of published preparation called Melzer’s reagent (blue, red– culture descriptions currently available (see Bibliog- brown, or non-reactive). raphy). Under these circumstances, one way to iden- The character and appearance of the spores and tify an unknown culture is to match it with others various elements such as cystidia and metuloids that isolated from the tissues of verified fruitbodies. may be present in the hymenium layer are also impor- There are also several compatibility tests that can be tant features for distinguishing agaric species, as is employed, such as the Buller test. This makes use of the internal arrangement and structure of the hyphae the observation that when an unknown field isolate is within the gills or lamellae. paired in culture with single-spore test isolates of known identity, a positive interaction will occur only

Figure 3. An assortment of microscopic elements from basidiomycete fruitbodies. The left half of the figure is composed of hyphae: (A) generative hyphae, with and without clamp connections at the septa are situated top left, and (B) two skeletal hyphae and a (C) branched binding hypha are at the lower left. At the centre and top right are sections of two hymenial (spore-bearing) surfaces with a (D) non-septate basidium (and attached spores) and a pointed (E) seta (centre), and on the right (left-to-right) are a specialised (F) setoid element with spines, a (G) cystidiole and a (H) crystal- encrusted lamprocystidium (metuloid). Various basidiospores (I) belonging to different species are present lower right.

39 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. with isolates of the same species, thus allowing a suc- which also enables identification if the identity of one cessful identification of the field isolate. A positive of the cultures is known. However, this method has interaction is indicated by a change in the nature of its limitations, and a control pairing using a different the test isolate from the haploid to dikaryotic condi- genotype of the same species should form part of the tion (cells with two nuclei, n + n, Figure 2) as a result procedure to demonstrate the appearance of a clear of the movement of nuclei from the field culture. This zone of incompatibility. change can be detected by microscopic examination The rapid development of molecular technology (e.g. by the appearance of hyphae with clamps at has provided another approach which is already septa within the test isolate, which occurs only when being used to identify unknown basidiomycete iso- the hyphae become dikaryotic), and sometimes by lates, by comparing DNA profiles with those of cul- changes in the visible appearance of the test culture tures from authenticated fruitbodies. As with cultural itself. Again, the uniform merging of two dikaryotic methods, which are limited by a lack of available cultures without the formation of an incompatibility culture descriptions, it is still necessary to build up barrier zone may indicate that the cultures belong to reference libraries of DNA profiles of accurately the same genotype and therefore to the same species, identified species.

Figure 4. A collage of micro-elements from basidiomycete cultures. Straddling a crystal-encrusted lamprocystidium (A – centre) are three (B) thick-walled, non-septate skeletal hyphae (branched and unbranched; binding hyphae are not distinguished in cultural descriptions). To the left are (C) clamped generative hyphae and a hypha with (D) encrustations of crystals; (E) a hypha with two clamps at the septum is on the right. At the top right are vegetative propagatory microstructures: (F) oidia (arthrospores), above, and a (G) thick-walled, intercalary chlamydospore. At lower right is a (H) ‘hyphal knot’, (I) a jigsaw-like tissue of interlocking hyphae, and a cluster of (J) rounded cuticular cells.

40 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

The procedures for describing basidiomycete surface bright orange–brown, eventually paling, species in culture rely on features such as the growth pores fine (6–9 per mm). In section, context pale col- rate on standard medium, colony colour, appearance, oured. Monomitic, generative hyphae thin- or thick- odour, and colour reaction to a selection of reagents. walled, with cross walls (septa), without clamps, Microscopic characters include the presence or absence hyaline (colourless). Hymenium with cystidioles of clamps at the colony margin and in the older culture, (Figure 3), basidiospores sub-globose, thin-walled, various types of hyphae and propagules, and a number colourless, smooth, non-staining in Melzer’s reagent of other distinctive microstructures (Figure 4). (inamyloid), 3.5–4.5 × 3.5–4 µm. Unlike the related R. lineatus, has no encrusted metuloids (lampro- Some basidiomycetes causing root cystidia) in either fruitbody or culture. Causes a root disease of planted trees (e.g. rubber, Hevea); produces disease in tropical hardwood white, branching mycelial cords and a white rot. plantations Junghuhnia vincta (synonyms, Poria vincta, The following basidiomycetes, all in the Aphyllopho- Rigidoporus vinctus) (Figure 5) rales, cause significant root disease or heart rot in tropical forests and plantations. All are known in Indonesia. Fruitbody a pinkish (variety vincta ) or greyish- Brief descriptions are given to emphasise key macro- brown (variety cinerea) resupinate crust; pores fine scopic and microscopic distinguishing features. Unfor- (6–12 per mm). Dimitic, generative hyphae thin- tunately, fruitbodies may not appear on infected trees walled, with cross walls (septa), without clamps, until disease is well advanced, so identification gener- hyaline (colourless); skeletal hyphae thick-walled, ally relies on field symptoms or the isolation of charac- slightly coloured, without septa. Hymenium with teristic cultures. Many basidiomycete species have cystidioles and encrusted metuloids; basidiospores changed their names over the years as more has been sub-globose, thin-walled, colourless, smooth, non- learned about their relationships, so some older syno- staining in Melzer’s reagent (inamyloid), 3.5–5.5 × nyms are also provided to help avoid confusion. 3–4 µm. Occasional clamps in culture. May fruit in older culture. Causes a root disease of planted trees. Rigidoporus microporus (synonyms, Produces a white rot. R. lignosus, Fomes lignosus) Phellinus noxius (Figure 6) Fruitbody a broad (to 20 cm wide), relatively thin, annual to less frequently perennial, leathery, broadly Fruitbody a cinnamon-brown, broadly attached, attached shelf, clustered, often imbricate; upper sur- woody shelf, developing a blackish crust on the upper face, concentrically furrowed, initially orange–red– surface, or a similar coloured resupinate sheet. Lower brown, faintly velvety, later smooth, faded; lower surface, greyish to dark brown, pores fine (6–8

Figure 5. Juhnghuhnia vincta. Left-to-right, poroid, fruitbody crust; section of fruitbody showing pore zone; microscopic elements of fruitbody, including a (A) sectioned metuloid, (B) hymenium zone with basidium, spores, and two cystidioles; (C) two thin-walled hyphae with septa and (D) three thick- walled, non-septate skeletal hyphae. After Hood (2003)

41 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. per mm). Tissues golden-brown within. Dimitic, with binding hyphae. The skeletal hyphae are the most narrow generative hyphae having septa that lack obvious within the tissues, and clamped generative clamps, and wider, thick-walled, non-septate, golden- hyphae are not easily seen. Spores double-walled, yellow skeletal hyphae. No setae (Figure 3) are present with fine spines between the two layers, apex trun- in the hymenium (as in some other Phellinus species), cated, ovoid, pale brownish, 6–9.5 × 4–8 µm. but characteristic long, thick-walled, pointed, non-sep- Causing root disease in hardwood hosts (e.g. rubber, tate, deep red–brown setal hyphae (tramal setae) are cacao, tea). Infected roots are coated in a reddish or found within the tissues. Spores smooth, ovoid, colour- reddish-brown mycelial crust which has a creamy- less, 3.5–5 × 3–4 µm. Fruitbodies are not always seen, white rhizomorph-like growing margin. Produces a but infected trees are recognised by a characteristic white rot. brown mycelial weft (with setal hyphae present) or a Ganoderma philippii belongs to the Ganoderma- black mycelial crust that forms on infected roots and taceae, a family characterised by a dimitic or trimitic develops collar-like around the base of the stem. hyphal system with clamped generative hyphae Decayed wood eventually becomes pale and honey- (often difficult to find), and especially by a character- combed in sheets, with brown zone lines. istic spore with tiny spines or echinulae positioned Phellinus noxius belongs to the Hymenochaetaceae, between two walls. The family includes Amauro- a family characterised by fungi with cinnamon-brown derma, with species that have stalked fruitbodies, and or golden yellow tissues, and generative hyphae Ganoderma, which forms annual or perennial, corky without clamps. Phellinus species are all dimitic. Most or woody brackets. Ganoderma species are divided do not cause disease, but many are responsible for into two groups, those with a non-shiny upper fruit- heart rots in living trees on which they produce prom- body surface formed of interwoven hyphae inent shelves or hoof-like brackets. (e.g. G. philippii, G. australe, Figure 7, upper left), and others, often stalked, with a shiny, lacquered Ganoderma philippii (synonym, surface composed of a palisade of hyphal ends G. pseudoferreum) (Figure 7) (Figure 7, centre; e.g. G. cupreum, G. steyaertanum, Figure 7, centre, upper right). Ganoderma species are Fruitbody a broadly attached, woody shelf, con- highly variable and difficult to separate into reliable centrically furrowed and warty above, smooth, semi- taxa using conventional morphology. While some glossy in parts, coloured dark reddish- or purplish- species in both groups are well understood, other brown, with a narrow, white margin; white or even- ‘species’ can still not be reliably resolved, and many tually brownish, beneath, with medium-fine pores laccate forms will probably prove to be synonyms of ((4–)6–7 per mm). Tissues brown. Often grouped, other taxa. Under these circumstances, the precise imbricate. Trimitic, with narrow generative hyphae, identification of the species responsible for root having septa with clamps; wider, thick-walled, non- disease in Acacia mangium plantations is a consider- septate, brown skeletal hyphae that branch near the able challenge. The genus Ganoderma is currently ends; and narrower, thick-walled, brown, branching ‘in taxonomic chaos’ (Ryvarden 1991, 1995), but it

Figure 6. Phellinus noxius. Fruitbody (right) and segment of crust (left) taken from a diseased root. Centre, microscopic features, including (top to bottom) (A) dark-brown rounded cells from mycelial crust, (B) skeletal hyphae, (C) two thin-walled, septate, clamp-less, generative hyphae, and (D) two thick- walled, setal hyphae with pointed ends. After Hood (2003)

42 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. appears that the use of molecular technology will G. australe, G. cupreum), while others are harmful soon result in a more satisfactory situation. Some parasites (e.g. G. philippii, G. steyaertanum and the species of Ganoderma are saprobes, causing heart European laccate species, G. lucidum). rots in living trees and decaying fallen wood (e.g.

Figure 7. Ganoderma species. Fruitbodies of G. australe, with a dull upper surface (top left), and G. steyaertanum (top right) and G. cupreum (centre left), both with a shiny red laccate surface, turning into a black crust in the case of the latter. Section of Ganoderma sp. fruitbody (lower left). Microscopic features: section of upper surface of fruitbody with a laccate crust (centre right), and distinctive spore and branched ends of two skeletal hyphae (lower right). All three species are reported from Indonesia and elsewhere. After Hood (2003)

43 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

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44 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Basidiomyctes – semi-popular Sen, M. 1973. Cultural diagnosis of Indian Polyporaceae. 3. Genera Daedalea, Favolus, Ganoderma, Hexagonia, Hood, I.A. 2003. An introduction to fungi on wood in Irpex, Lenzites, Merulius and Poria. Indian Forest Queensland. Armidale, Australia, University of New Records (new series), Forest Pathology, 2(11), 277–304 England, School of Environmental Sciences and Natural (Plates I–XI). Resource Management, 388p. Stalpers, J.A. 1978. Identification of wood-inhabiting Zoberi, M.H. 1972. Tropical macrofungi, some common Aphyllophorales in pure culture. Studies in Mycology, species. London, Basingstoke, Macmillan, 158p. 16, 1–248. Baarn, Netherlands, Centraalbureau voor Schimmelcultures. Identification in culture Van der Westhuizen, G.C.A. 1971. Cultural characters and carpophore construction of some poroid hymenomycetes. Bakshi, B.K., Sehgal, H.S. and Singh, B. 1969. Cultural Bothalia, 10(2), 137–328. diagnosis of Indian Polyporaceae. 1. Genus Polyporus. Indian Forest Records (new series), Forest Pathology, 2(9), 205–244 (Plates I–XIII). Identification from field symptoms and signs Bakshi, B.K., Sen, M. and Singh, B. 1970. Cultural Flood, J.; Bridge, P. and Holderness, M., ed. 2000. diagnosis of Indian Polyporaceae. 2. Genera Fomes and Ganoderma diseases of perennial crops. Wallingford, Trametes. Indian Forest Records (new series), Forest UK, CABI International, 288p. Pathology, 2(10), 245–275 (Plates I–XI). Gadgil, P.D. 2005. Fungi on trees and shrubs in New Nakasone, K.K. 1990. Cultural studies and identification of Zealand. Fungi of New Zealand, Volume 4, Fungal wood-inhabiting Corticiaceae and selected Diversity Research Series 16. Hong Kong, Fungal Hymenomycetes from North America. Mycological Diversity Press, 437p. Memoir, 15, 1–412. Old, K.M., Lee, S.S., Sharma, J.K. and Yuan, Z.Q. 2000. A Nobles, M.K. 1948. Identification of cultures of wood- manual of diseases of tropical acacias in Australia, South- rotting fungi. Canadian Journal of Research, 26, 281– East Asia and India. Jakarta, Indonesia, Centre for 431. International Forestry Research, 104p. — 1965. Identification of cultures of wood-inhabiting Sripathi Rao, B. 1975. Maladies of Hevea in Malaysia. hymenomycetes. Canadian Journal of Botany, 43, 1097– Kuala Lumpur, Rubber Research Institute of Malaysia, 1139. 108p.

45 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

The use of DNA techniques to identify fungi

Morag Glen1

Abstract

DNA provides an abundance of taxonomic characters for the identification of organisms that have inadequate morphological characters, or possess distinguishing features only during particular stages of their life cycle. Methods of accessing these DNA characteristics vary, though variations of PCR-based techniques are popular at present, because of their speed, sensitivity and high throughput capability. Several popular methods are described that have been applied in diverse areas including food quality, forest ecology, and human, animal and plant pathology. The method selected for a particular application will depend on several factors, including the number of samples and number of candidate species. All DNA-based techniques depend on an adequate herbarium resource of carefully preserved specimens with detailed morphological descriptions to verify the DNA-based protocol.

DNA techniques are a valuable tool in the identifica- fungi in preserved and manufactured foods and tion of fungi. They are particularly useful as a means species verification of high-value truffles. to determine the identity of a fungus when it is not DNA identification of plant-associated fungi also producing fruitbodies, the morphological taxonomic has many applications, including community studies characters that form the basis of the species descrip- of beneficial mycorrhizal fungi, monitoring the con- tion. While some fungi can be grown in culture with tinued presence of inoculated biocontrol or symbiotic ease and produce characteristic microscopic features fungi, and early detection of disease-causing fungi to that assist in taxonomic identification, others are dif- allow timely management. ficult to grow in culture or, when grown in culture, fail to develop key distinguishing features. The need for rapid identification, as in quarantine or biosecu- DNA basics rity applications, may also justify the use of DNA DNA (deoxyribonucleic acid) is a long molecule techniques. constructed from a variable number of nucleotides, DNA techniques for the identification of fungi also called bases or base-pairs (bp). Each nucleotide have been widely used in human and veterinary med- consists of a 5-carbon sugar molecule, deoxyribose, icine, where rapid and accurate identification of with a phosphate group attached to carbon 5 and one fungal pathogens assists the selection of appropriate of four bases attached to carbon 1 (Figure 1). The treatment. They have also been applied to food four bases are adenine, guanine, cytosine and quality control for the detection of contaminating thymine, commonly referred to as A, G, C and T, respectively. As a strand of DNA is synthesised, the 1 Ensis – the joint forces of CSIRO and Scion, Private Bag phosphate group of a nucleotide is chemically 12, Hobart, Tasmania 7001, Australia. bonded to the 3' hydroxyl group of the last nucleotide Email: . in the growing chain.

46 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

the number of samples. However, the foundation for all methods is the use of a DNA ‘signature’ to link the unknown fungus to a fully described, morphologically characterised herbarium specimen. Identifica- tion of a broad range of fungi is therefore dependent on a comprehensive set of reference specimens that have been examined and identified by a mycological taxonomist with the appropriate skills for the class of fungi under consideration. Public DNA sequence databases can be used as additional sources of infor- Figure 1. Deoxyadenosine triphosphate (dATP) one mation, but should not be relied upon except for ten- of the four deoxynucleotides (dNTPs) tative identifications, as validating the identity of needed for synthesis of DNA source material is rarely possible. Methods for DNA identification of fungi include: DNA occurs in cells as a double-stranded molecule, Southern blotting; PCR (polymerase chain reaction); held together by electrostatic forces between the bases PCR-RFLP (PCR restriction fragment length poly- in strand 1 and those in strand 2. Guanine is electrostat- morphism); T-RFLP (terminal restriction fragment ically attracted to cytosine, and adenine to thymine, and length polymorphism); DNA sequencing; and micro- these pairs are called ‘complementary’. The double arrays. The first step for any of these methods is DNA strand is thermodynamically stable under physiological extraction and purification, for which many protocols conditions if the two strands consist of complementary and manufactured kits are available. base sequences (Figure 2). DNA polymerase is an enzyme that synthesises DNA by copying an existing Southern blotting strand of DNA. It does not make an exact copy, but a ‘reverse complement’. The two strands of DNA are Southern blotting or dot blotting involves binding separated and a ‘reverse complement’ made of each genomic DNA to a membrane, denaturation of the strand, so that strand 1 is the template for a new strand bound DNA, then hybridisation with a labelled probe 2, and vice versa. It is the order or sequence of the As, (Goodwin et al. 1989; Figure 3). The probe consists of Cs, Gs and Ts that provides the genetic information that a short fragment (20 or more bp) of DNA that is unique is passed from cell to cell and generation to generation, to the target organism. Because of the requirement for and it is determining this sequence of nucleotides that is comparatively large quantities of DNA, this method referred to as ‘DNA sequencing’. has largely been superseded by PCR-based methods. PCR Methods PCR was developed in the 1980s and exploits a There are several techniques that can be used for thermostable DNA polymerase enzyme from a ther- fungal identification. The selection of the most motolerant bacterium so that the enzyme can with- appropriate method depends on the application and stand the temperature cycling required for PCR

Figure 2. Two DNA strands with complementary base pairs form a stable duplex (A), but two strands with a high number of mismatches do not (B)

47 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

(Saiki et al. 1988). PCR is used to make many copies is achieved by heating the reaction mixture. It is then of a small portion, usually up to 1 kbp, of genomic cooled to promote primer annealing and heated again DNA. Two primers, oligonucleotides of approxi- to reach the optimum temperature for the DNA mately 20 bases, with base sequences complemen- polymerase to extend the primers, making a reverse tary to a small fragment on either side of the target complement copy of the template DNA strand. This DNA are also required to initiate replication. Separa- three-step heating and cooling profile is repeated for tion of the two template DNA strands is necessary 30–40 cycles, allowing a theoretical doubling of the before the primers can bind to the template, and this target sequence with each cycle (Figures 4 and 5).

Figure 3. A. Southern blotting procedure. B. Dot blotting does not require gel electrophoresis and transfer, instead the DNA solution is spotted directly onto the membrane. DNA is denatured by heat or alkali, to separate the two strands, then applied to a nylon or nitrocellulose membrane in a regular pattern by pipette or by suction through a special device. The membrane is then incubated in a solution containing labelled probe DNA, which binds only to DNA of the target species. The label can be radioactive, fluorescent or enzymatic. After development of the radiogram (for radioactive probes), the DNA spot from the target species will produce a dark spot on the film.

48 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

During a 30-cycle PCR, assuming 100% efficiency, modulin (Mulè et al. 2004) or multi-copy regions 230 (i.e. 109) copies of a target sequence can be made, such as rDNA ITS (ribosomal DNA internal tran- sufficient for visualisation on ethidium bromide scribed spacers) for greater sensitivity (Flowers et al. stained gels. 2003). Extensive testing is required to validate the To use PCR as a fungal identification tool, species- species-specificity of the PCR test. Internal amplifi- specific primers are required, which will amplify a cation controls are also necessary to provide confi- product from only the target species. Species-specific dence in negative results (Hoorfar et al. 2004). primers can be designed for well-characterised genes Species-specific PCR can be adapted to high- such as calmodulin (Mulè et al. 2004) or unknown throughput equipment, allowing the rapid processing sequences such as RAPD (random amplified poly- of many samples, but the method is suitable for iden- morphic DNA) or AFLP (amplified fragment length tification of only one or a few fungi, as each sample polymorphism) products (Dobrowolski and O’Brien must be tested for each species of interest. Applica- 1993; Vos et al. 1995; Schmidt et al. 2004). Primers tions include early detection and identification of can be made to target a single-copy gene such as cal- plant pathogens, monitoring the spread of pathogens

Double-stranded DNA template

Primers dNTPS polymerase enzyme a) PCR ingredients

b) Step 1: 95°C, DNA denatures and the two strands separate

c) Step 2: 45–65°C, Primers anneal to complementary sequences

d) Step 3: 72°C, Polymerase extends primers, adding dNTPs to growing oligonucleotide chain

e) At the end of cycle 1, there can be twice as many copies of the target region

Figure 4. A schematic representation of cycle 1 of a PCR

49 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. or biocontrol organisms and validation of disease- PCR product with an enzyme that has a 4bp recogni- free status for safe germplasm movement. tion site will typically cut the PCR product into 2–8 fragments of assorted sizes that will generate a par- PCR-RFLP ticular profile when subjected to electrophoresis on PCR-RFLP starts with a PCR, using primers with an agarose or acrylamide gel (Figure 6). This method broad specificity, such as primers ITS1-F and ITS4, has been widely used in ectomycorrhizal research as that will amplify a product from most if not all fungi, it is suitable for samples that contain mainly one but do not amplify plant, animal or bacterial DNA. fungus that could belong to any of a large number of The PCR product is digested with a restriction species (Gardes and Bruns 1993). Digestion with two enzyme that cuts the DNA strands at a particular rec- or three different restriction enzymes is usually ognition site for each enzyme. The recognition site required to distinguish most species. While many for Hae III, for example, is GGCC. Cutting a 1 kbp species can be clearly distinguished using this

Figure 5. A representation of the first three cycles of a PCR, showing the production of specific fragments delimited by primer sequences.

50 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. method, some genera such as Tomentella and Corti- herbarium specimens. The more extensive the refer- narius have many species with very similar profiles ence database, the greater the likelihood of obtaining that are difficult or impossible to distinguish. a match, and the lower the risk of applying an incor- A sizing error of up to 5% must be accommodated in rect name. Profiles must be 100% identical — a profile matching, rendering highly similar profiles dif- partial match does not provide useful information for ficult to distinguish. In addition, in using this technique, either PCR-RFLP or T-RFLP. there are often many DNA profiles that are not matched with known species, though these unmatched fungi can DNA sequencing still be monitored for prevalence and abundance. DNA sequencing often begins with a PCR to provide If warranted, further information can be sought by sufficient template DNA for the sequencing reaction, DNA sequencing (see below). PCR-RFLP has also though this may also be achieved by cloning the been used in identification of fungal pathogens desired fragment into a plasmid or bacteriophage. (Rolshausen et al. 2004), but the presence of multiple The sequencing reaction is very similar to a PCR, fungal species in one sample can render the PCR-RFLP though only one primer is used per reaction and, in profile very difficult to interpret. addition to the deoxynucleotides, di-deoxynucleotides (ddNTPs) are included in the reaction mix. T-RFLP These ddNTPs are labelled with one of four fluores- T-RFLP is a variant of PCR-RFLP that uses a flu- cent dyes corresponding to each of the four bases, A, orescent primer and a sequencing gel to obtain G, C and T. When a ddNTP is incorporated into the greater sensitivity and fragment size accuracy. How- newly synthesised molecule, growth of this chain ever, only the fragment with the fluorescent primer stops as the ddNTP lacks the oxygen molecule to registers on the sequencing gel, so the profile for each which the next dNTP would be attached. The final fungus/enzyme combination is a single fragment. product is therefore a series of fragments ranging in This method has been used mainly in ectomycor- size from ~20 bp to n bp, where n is the length of the rhizal research (Dickie et al. 2002). Both PCR-RFLP PCR product used for template (Figure 7), each frag- and T-RFLP require the establishment of a reference ment having one fluorescently labelled, terminal database of fragment sizes from identified fungi with ddNTP. Electrophoresis on a long, high-resolution

Figure 6. Agarose gel of PCR-RFLP products. Lanes 1, 15 and 30 contain DNA size markers. These can be used to estimate the sizes of the PCR-RFLP fragments. The patterns from known species can be stored as a string of fragment sizes in a searchable database. Usually digestion with two different enzymes will provide adequate discrimination among species, though certain genera with many closely related species may be more difficult.

51 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. acrylamide gel or capillary separates DNA fragments sometimes with the assistance of phylogenetic analysis that differ in length by as little as one nucleotide. As (Bruns et al. 1998; Glen et al. 2002). the fragments move past a scanner, shorter fragments This would identify target genera for expansion of followed by fragments of increasing length, the fluo- the reference database. This method of identification is rescent tag on the ddNTP ‘calls’ the base sequence applicable to many areas of research, and is particularly (Figure 8), which is converted by the software to a suitable where there is a large number of candidate string of As, Cs, Gs and Ts. species and it is known that not all species of interest For fungal identification, a database of sequences are included in the reference database. The occurrence from identified fungi is searched with the sequence of more than one fungal species in a sample, such as from the unknown fungus. If a 100% match is not may occur in a well-rotted root, can cause problems, found, information can be derived from the best per- though, if warranted, cloning of the PCR product centage match and a likely genus may be inferred, before sequencing can accommodate these difficulties.

Figure 7. During a dye terminator sequencing reaction, a single primer is used to initiate copies of one of the two strands of DNA template. Each copy is terminated at a random point by the incorporation of a fluorescently labelled dideoxy nucleotide (ddNTP). The products therefore consist of single stranded DNA of varying length.

52 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Micro-arrays (Anderson et al. 2005). This technique is expensive to develop but will be well suited to analyses A promising new approach is the use of DNA requiring very high throughput. micro-arrays for fungal identification. DNA microarrays exploit probe/target binding in a similar manner to Southern or dot blotting, though the format Application to heart and root rot in is much smaller and the support consists of a glass Acacia mangium slide rather than a nylon or nitrocellulose membrane (Southern 1996). Many probes can be crammed into Many fungi are implicated in heart and root rots in a very small area, making this method ideal for high A. mangium. Currently, the most suitable method of throughput or simultaneous analysis of many genes detecting/identifying these fungi involves the use of — hence the popularity of the micro-array in gene PCR and DNA sequencing, as species-specific expression studies examining a large number of primers have been developed for only a few of the genes (Ramsay 1998). This technique has been used many root or butt rot species (Lim et al. 2005; Suhara for identification of medically significant fungi et al. 2005). While many sequences of fungi of (Leinberger et al. 2005) and work is proceeding on interest, such as those of Ganoderma spp. and Phell- developing the method for high throughput detection inus spp., are available on public DNA databases, not and identification of plant pathogenic fungi many of them are linked to herbarium specimens that

Figure 8. The products of the dye terminator sequencing reaction are subjected to electrophoresis on a high-resolution gel or capillary that passes a scanner attached to a computer. The shorter fragments migrate faster through the gel and therefore reach the scanner first, followed by the longer fragments. The scanner picks up the signal from each fluorescent nucleotide as it passes, and this is converted by the computer to an electropherogram, above, and a DNA base sequence. The electropherogram provides visual confirmation of data quality, allowing manual editing of the derived sequence.

53 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. can be readily accessed. Accumulating a comprehen- Hoorfar, J., Cook, N., Malorny, B., Wagner, M., De Medici, sive database of fungal sequences linked to her- D., Abdulmawjood, A. and Fach, P. 2004. Diagnostic barium vouchers with detailed macroscopic and PCR: making internal amplification control mandatory. microscopic descriptions is therefore a high priority. Applied Microbiology, 38, 79–80. The taxonomy of the polypore species associated Leinberger, D.M., Schumacher, U. Autenrieth, I.B. and with heart, butt or root rot is, in many cases, in need Bachmann, T.T. 2005. Development of a DNA of clarification and DNA sequencing can help microarray for detection and identification of fungal resolve some species delimitations. pathogens involved in invasive mycoses. Journal of Clinical Microbiology, 43, 4943–4953. Lim, Y.W., Yeung, Y.C.A., Sturrock, R., Leal, I. and References Breuil, C. 2005. Differentiating the two closely related species, Phellinus weirii and P. sulphurascens. Forest Anderson, N., Szemes, M., de Weerdt, M., Vahlkamp, L., Pathology, 35, 305–314. Boender, P., Schoen, C., Bonants, P. and O’Brien, P.A. Mulè, G., Susca, A., Stea, G. and Moretti, A. 2004. Specific 2005. Multiplex detection of plant pathogens by detection of the toxigenic species Fusarium proliferatum microarrays: an innovative tool for plant health and F. oxysporum from asparagus plants using primers management. 15th Biennial Conference of the based on calmodulin gene sequences. FEMS Australasian Plant Pathology Society, Conference Microbiology Letters, 230, 235–240. Handbook, 74 p. Bruns, T. D, Szaro, T. M, Gardes, M., Cullings, K. W, Pan, Ramsay G. 1998. DNA chips: state-of-the-art. Nature J. J, Taylor, D. L, Horton, T. R, Kretzer, A., Garbelotto, Biotechnology, 16, 40–44. M. and Li, Y. 1998. A sequence database for the Rolshausen, P.E., Trouillas, F. and Gubler, W.D. 2004. identification of ectomycorrhizal basidiomycetes by Identification of Eutypa lata by PCR-RFLP. Plant phylogenetic analysis. Molecular Ecology, 7, 257–272. Disease, 88, 925–929. Dickie I. A., Xu B. and Koide R. T. 2002. Vertical niche Saiki, R.K., Gelfand, D.H., Stoffel, S., Scharf, S.J., Higuchi, differentiation of ectomycorrhizal hyphae in soil as R., Horn, G.T., Mullis, K.B. and Erlich, H.A. 1988. shown by T-RFLP analysis. New Phytologist, 156, 527– Primer-directed enzymatic amplification of DNA with a 535. thermostable DNA polymerase. Science, 239, 487–491. Dobrowolski, M. P and O'Brien, P. A 1993. Use of RAPD- Schmidt, H., Taniwaki, M.H., Vogel, R.F. and Niessen, L. PCR to isolate a species specific DNA probe for 2004. Utilization of AFLP markers for PCR-based Phytophthora cinnamomi. FEMS Microbiology Letters, identification of Aspergillus carbonarius and indication 113, 43–48. of its presence in green coffee samples. Journal of Flowers, J., Hartman, J. and Vaillancourt, L. 2003. Applied Microbiology, 97, 899–909. Detection of latent Sphaeropsis sapinea infections in Southern, G.M. 1996. DNA chips: analysing sequence by Austrian pine tissues using nested-polymerase chain hybridization to oligonucleotides on a large scale. Trends reaction. Phytopathology, 93, 1471–1477. in Genetics, 12, 110–115. Gardes, M. and Bruns, T.D. 1996. Community structure of Suhara, H., Maekawa, N., Kubayashi, T. and Kondo, R. ectomycorrhizal fungi in a Pinus muricata forest: above- 2005. Specific detection of a basidiomycete, Phlebia and below-ground views. Canadian Journal of Botany, brevispora associated with butt rot of Chamaecyparis 74, 1572–1583. obtusa, by PCR-based analysis. Journal of Wood Science, Glen, M., Tommerup, I.C., Bougher, N.L., O’Brien, P.A. 51, 83–88. 2002. Are Sebacinaceae common and widespread Vos, P., Hogers, R., Bleeker, M., Reijans, M., Lee, T. van de, ectomycorrhizal associates of Eucalyptus species in Hornes, M., Frijters, A., Pot, J., Peleman, J., Kuiper, M. Australian forests? Mycorrhiza, 12, 243–247. and Zabeau, M. 1995. AFLP: a new technique for DNA Goodwin, P. H, Kirkpatrick, B. C and Duniway, J. M 1989. fingerprinting. Nucleic Acids Research, 23, 4407–4414. Cloned DNA probes for identification of Phytophthora parasitica. Phytopathology, 79, 716–721.

54 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Molecular identification of organisms associated with root and heart rot in Acacia mangium

Morag Glen1, Karina Potter1,2 and Purnamila Sulistyawati3

Abstract This paper discusses the application of molecular identification of fungi to research into heart and root rots of Acacia mangium, using four different examples. These examples illustrate the utility of DNA sequence information in clarification of the ecology, biology and pathology of fungal species

An overview of different techniques for molecular databases. This may give identification only to genus identification of fungi is given by another paper in or family level in some cases, but this level of infor- these proceedings (Glen 2006). All of these tech- mation allows the selection of specific groups of niques are based on linking an unknown organism to fungi for further analysis. a well-characterised herbarium specimen by a similar The ribosomal DNA internal transcribed spacers DNA profile. As a large number of fungal species are (rDNA ITS) is a genomic region that is widely used implicated in heart and root rots, a technique that can in fungal systematics and identification (Figure 1). simultaneously identify many species is more effi- Its advantages lie in the highly conserved nature of cient than species-specific probes or primers. In addi- the ribosomal DNA genes, which means that primers tion, as no comprehensive database of root and heart can be designed to amplify DNA from a broad range rot fungi from Acacia mangium Willd. exists, tech- of species and the high degree of variation in the non- niques that require an exact match, such as PCR- coding region. As the spacers region does not code RFLP and T-RFLP, will provide only limited infor- for a functional gene, there is little, if any, constraint mation. on mutations in this region. There is, therefore, a The method that is most likely to efficiently useful degree of inter-specific variation that facili- provide a useful level of information is to sequence a tates species discrimination and identification. In well-characterised region of DNA and compare this addition, the ribosomal DNA genes are contained in to sequences of known fungi from public and private tandem repeats, with up to 50 copies per genome. This makes amplification by PCR much easier than 1 Ensis – the joint forces of CSIRO and Scion, Private Bag for a single-copy gene. 12, Hobart, Tasmania 7001, Australia. DNA sequencing has been used to identify fungi Email: . 2 School of Agricultural Science, University of Tasmania, causing disease in A. mangium in four different Private Bag 12, Hobart, Tasmania 7001, Australia. experiments, as part of the ACIAR ‘Heartrots in Email: . plantation hardwoods’ project (FST/2000/123): 3 Ministry of Forestry FORDA, Centre for Plantation Forest Research and Development, Jalan Palagan 1. Detection of Ganoderma and Amauroderma Tentara Pelajar Km. 15, Purwobinangun – Pakem, mycelium in rotten root and butt samples from Yogyakarta 55585, Indonesia. trees with and without associated fruitbodies. Email: . 2. Fungi isolated from a provenance wounding trial.

55 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Figure 1. A schematic representation of the organisation of the rRNA gene repeat indicating the location of the internal transcribed spacers (ITS). B gives an enlarged view of the circled section in A. Primer sites for DNA amplification from a broad range of fungi are located in the 18S and 25S rRNA genes. 3. Direct detection of fungi in woodblocks from the the wounds were a Pycnoporus sp. and an unidenti- same wounding trial. fied fungus, heart-rot isolate 3-2-A1. The third 4. Identification of Ganoderma spp. isolates wound was left uninoculated. The 72 trees were recovered from roots used in somatic felled 11 months after inoculation and the wood was incompatibility tests. assessed for fungal decay and discoloration. An In the first experiment, samples of rotten roots and attempt was made to isolate fungi from three regions butts of A. mangium trees that showed symptoms of around each of the three wounds per tree (Figure 3). ‘red root rot’ were collected, along with fruitbodies of Some 172 fungal isolates were obtained, but none of Ganoderma philippii, Amauroderma rugosum and the isolates recovered from the experiment corre- other fungal species from several A. mangium planta- sponded morphologically to the isolates used in the tions throughout Indonesia (Glen et al. 2006). Gano- wound inoculation. derma philippii was suspected to be the causal agent of The recovered isolates were sorted into 15 groups this red root-rot disease, based on similarities to disease based on gross morphological characteristics. caused by this species in other woody crops and the Enzyme production from these isolates was tested prevalence of G. philippii fruitbodies in affected plan- using spot tests. Only one of the morphological tations. The rDNA ITS was amplified and sequenced groups appeared to produce easily detectable quanti- from the fruitbodies. Fungal rDNA was also amplified ties of laccase, indicating white-rot capabilities from the root and butt samples using fungi-specific (group 7). Low levels of tyrosinase were also primers to avoid the amplification of DNA from bac- detected in this group. The ITS of a sub-set of isolates teria, insects or plants that may have been present in the (n = 20) were sequenced, focusing primarily on sample. The PCR products amplified from the rot group 7. Results of BLAST searches of public data- samples were clean enough to sequence directly, indi- bases are given (Table 1). A sequence has not yet cating that the samples were colonised by one predom- been successfully obtained from either of the original inant fungus. The sequences matched those from the G. fungal isolations used to inoculate the wounds. philippii fruitbodies (Figure 2). This indicated the pres- The first three fungi in Table 1 (Oudemansiella, ence of the fungus in the rotten wood, though was Pycnoporus and Trametes) are likely to have a role in insufficient evidence to prove that the rot was caused heart rot, as enzyme tests demonstrated the presence by G. philippii. The roles of other Ganoderma and of laccase and tyrosinase. While all these isolates Amauroderma species also need further investigation. were placed in group 7 on the basis of their gross In the second experiment, fungi were isolated as culture morphology and enzyme test results, they part of a provenance wounding trial that took place in were classified as three different basidiomycete fungi Sumatra in 2002–2003 (Barry et al. 2006). Trees based on their ITS sequences. The Pycnoporus were inspected for signs of heart rot before holes isolate recovered from the wounding trial wood were drilled at three heights and inoculum plugs block is likely to be the same fungus that was inocu- inserted into two of the holes, which were then lated, as its sequence is a very high match to the fruit- sealed. The two isolates of fungi used for inoculating body from which the original isolate was obtained.

56 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Figure 2. Maximum parsimony tree of ITS1 and ITS2 sequences from Ganoderma and Amauroderma species and sequences derived directly from wood. Bootstrap values over 50% are given. Scale bar represents 0.1 expected nucleotide substitutions. Collections from Acacia mangium plantations are in bold; sequences amplified from root or butt wood are marked with an asterisk. From Glen et al. (2006)

57 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

The Pycnoporus was re-isolated from a wood block (Irianto et al. 2006). Ganoderma lucidum consists of sampled from within the rotten area, while the Oude- a complex of at least three and possibly six species mansiella was isolated from a wood block taken from (Moncalvo et al. 1995a; Hseu et al. 1996). adjacent to the rotten area in a different tree that had also been inoculated with the Pycnoporus isolate. The Trametes sp. isolate is also a recognised wood rotter and was recovered from within the rotten area of a control wound. It is likely that the Oudeman- siella and Trametes isolates either invaded via the treatment wound or were already present before the inoculation. The remaining fungi are all ascomycetes, and while some may be important in other diseases, they do not have wood-rotting capabilities. The woodblocks from the wounding trial were stored to attempt direct detection of fungi. DNA was extracted from a small number of these woodblocks, but the PCR product was not clean enough to sequence. The PCR products are currently being cloned to obtain clean DNA sequences. The final example involves the identification of Figure 3. A diagram of the wood discs after fungi isolated from roots and fruitbodies from an harvesting, the grey area represents the A. mangium provenance/family trial that has suffered rotten wood surrounding the wound. Wood severe root rot at Wonogiri. The purpose of the blocks were removed from three areas of experiment was to determine the main mode of dis- each disc — (1) from within the rotten persal of the predominant fungus. Many Ganoderma wood, (2) adjacent to the rotten wood, and fruitbodies occur on this site and have been identified (3) away from the rotten section. as species from the Ganoderma lucidum complex

Table 1. Tentative identification of fungi isolated from the wounding trial

Isolatea Group Results of sequence database search 13A2-Y 7 Oudemansiella aff. canariensis 70A1 7 Pycnoporus aff. sanguineus/cinnabarensis 49C1 7 Trametes sp. 53A1 1 Phaeoacremonium aff. rubrigenum 58A1 2 Articulospora sp. 16C1-Y 4 Ascomycete, unknown species 69B2 6 Pestalotiopsis spp. 63C1 8 Phaeoacremonium aff. rubrigenum 69C1-X 8 Phaeoacremonium aff. rubrigenum 57B1-Y 8 Bionectria spp. 37C1 8 Clonostachys/Nectria gliocladioides 26A1-X 8 Clonostachys/Nectria gliocladioides 1C1-X 13 Aspergillus sp. 63A1-X Un-grouped Pestalotiopsis spp. 69C1-Y Un-grouped Nectria gliocladioides/Clonostachys sp. 67C1 Un-grouped Clonostachys sp./Nectria gliocladioides a The number before the letter in the isolate number indicates the tree number. The letter within the isolate number indicates the wound inoculum (A = Pycnoporus sp., B = heart-rot isolate 3-2-A1 and C = control, sterile inoculum) and the number after the letter indicates the location of the wood block sample (see Figure 3). The letter after the hyphen distinguishes individual isolates recovered from the same wood block.

58 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Fungi were isolated from fruitbodies and from the Glen, M., Bougher, N.L., Nigg, S.Q., Lee, S.S., Irianto, R., roots of diseased trees. These isolates were then used Barry, K.M. and Mohammed, C.L. 2006. Molecular for somatic incompatibility tests to determine the differentiation of Ganoderma and Amauroderma species number and extent of genetic individuals. In order to and their role in root rot disease of Acacia mangium validate the test results, it was necessary to demon- plantations in Indonesia and Malaysia. Australasian Plant Pathology, in press. strate that the isolates belong to the same species. ITS Gottleib, A.M., Ferrer, E. and Wright, J.E. 2000. rDNA sequences of the isolates confirmed the morpholog- analyses as an aid to the taxonomy of species of ical identification (Irianto et al. 2006) as Ganoderma Ganoderma. Mycological Research 104, 1033–1045. aff. lucidum. Phylogenetic analysis may allow us to Gottleib, A.M. and Wright J.E. 1999. Taxonomy of determine in which species group of the Ganoderma Ganoderma from southern South America: subgenus lucidum complex this species belongs. Elfvingia. Mycological Research 103, 1289–1298. Identification of Ganoderma species can be a chal- Hseu, R.S., Wang, H.H., Wang, H.F. and Moncalvo, J.M. lenge for even an experienced mycological taxono- 1996. Differentiation and grouping of the Ganoderma mist (Moncalvo et al. 1995a,b; Gottlieb and Wright lucidum complex by Random Amplified Polymorphic 1999; Gottlieb et al. 2000; Smith and Sivasitham- DNA-PCR compared with grouping on the bass of param 2000, 2003). DNA sequences can assist taxon- internal transcribed spacer sequences. Applied and omists to infer species boundaries and inter-specific Environmental Microbiology, 62, 1354–1363. relationships, and can also be used as a ‘fingerprint’ Irianto, R., Barry, K., Hidayati, N., Ito, S., Fiani, A., or ‘barcode’ to identify fungal species in life stages Rimbawanto, A. and Mohammed, C. 2006. Incidence and spatial analysis of root rot of Acacia mangium in that lack the characters on which traditional classifi- Indonesia. Journal of Tropical Forest Science, 18, 87–95. cation is based. This fingerprint can also be used by Moncalvo, J.M., Wang, H.F. and Hseu, R.S. 1995a. Gene the non-specialist to assist in classification, but close phylogeny of the Ganoderma lucidum complex based on co-operation with an experienced fungal taxonomist, ribosomal DNA sequences. Comparison with traditional and retention of reference specimens, are still vital. taxonomic characters. Mycological Research, 99, 1489– 1499. References — 1995b. Phylogenetic relationships in Ganoderma inferred from the internal transcribed spacers and 25S Barry, K.M., Irianto, R.S.B., Tjahjono, B., Tarigan, M., ribosomal DNA sequences. Mycologia 87, 223–238. Agustini, L., Hardiyanto, E.B. and Mohammed, C.L. Smith, B.J. and Sivasithamparam, K. 2000. Internal 2006. Variation of heartrot and extractives with transcribed spacer ribosomal DNA sequence of five provenance of Acacia mangium. Forest Pathology, 38, species of Ganoderma from Australia. Mycological 183–197. Research, 104, 943–951. Glen, M. 2006. The use of DNA techniques to identify fungi. — 2003. Morphological studies of Ganoderma (Ganoder- In: Potter, K., Rimbawanto, A. and Beadle, C., ed., Heart mataceae) from the Australasian and Pacific regions. rot and root rot in tropical Acacia plantations. Canberra, Australian Systematic Botany, 16, 487–503. ACIAR Proceedings No. 124, 46–54. [These proceedings]

59 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Root rot in tree species other than Acacia

A. Mohd Farid1, S.S. Lee1, Z. Maziah2, H. Rosli1 and M. Norwati1

Abstract

Surveys of root disease were conducted in forest plantations of Azadirachta excelsa, Tectona grandis and Khaya ivorensis throughout Peninsular Malaysia. Two major root diseases were found, namely white root disease and brown root disease caused by Rigidoporus lignosus and Phellinus noxius, respectively. A destructive root disease of K. ivorensis caused by an unidentified fungus was found in the state of Negeri Sembilan. These diseases were observed to be closely associated with poor land preparation and areas with a previous history of root disease. Based on experience gained from the management of root-rot disease in rubber plantations, good land management, the construction of isolation trenches and the application of fungicides are suggested as valuable tools in the control of root-rot disease in forest tree plantations.

Root disease causes significant mortality in many fruit trees (Singh 1973; Wood and Lass 1985; Ann et forest plantations and is a common explanation for al. 2002). failure in the early phase of plantation development In Malaysia, interest in forest plantations boomed (Wingfield 1999). In particular, root disease is a in the 1990s, and several fast-growing species were major threat to plantation monocultures that have introduced. Among these were sentang (Azadirachta been established on land converted from natural excelsa (Jack) Jacobs), teak (Tectona grandis L.) and forest with poor land-clearing techniques (Lee 1993). khaya (Khaya ivorensis A. Chev.). These species Furthermore, the low species and genetic diversity were planted with little knowledge of potential pest and uniform age of plantations create conditions and disease threats. In 1997, the Forest Research favourable for the development and spread of root- Institute Malaysia (FRIM) began disease surveys disease pathogens. throughout Peninsular Malaysia to determine the In India, Indonesia, Malaysia and Thailand, root health status of the most common forest plantation rot has been identified as the most serious disease in species. Root-disease incidence was the main thrust plantations of tropical acacias (Old et al. 1997). Root of the surveys as root diseases have the potential to rot is also the most destructive disease of rubber trees cause high levels of tree mortality. and can kill the tree irrespective of age or health status, causing economic losses to the latex industry in many countries (Nandris et al. 1987a; Liyanage Materials and methods 1997; Semangun 2000: Guyot and Flari 2002). The Disease surveys disease has also been reported to aggressively kill Root-disease surveys were conducted randomly on 1 Forest Research Institute Malaysia, Kepong, 52109 34 forest plantations throughout Peninsular Malaysia Kuala Lumpur, Malaysia. Email: . from 1997 (Mohd Farid et al. 2005). The surveys 2 School of Biological Sciences, Universiti Sains Malaysia, focused on plantations of sentang, teak and khaya. Penang, Malaysia. Email: . Complete and random censuses were conducted in

60 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. small (<0.4 ha) and large-scale plantations (>0.4 ha), ously reported that teak trees 2 years old and above respectively. In large-scale plantations, plots of 20 × were frequently infected by root disease in Malaysia, 20 trees were randomly established and surveyed. In and Browne (1968) notes that brown root disease was both small and large-scale plantations, trees selected destructive on teak of 2–12 years old in forest plan- for sampling were inspected individually. Back- tations in some countries. In this study, infection of ground information on the establishment of the plan- teak trees was recorded from as early as 1 year after tations was documented, including planting planting. techniques, land clearing, site history, source of Root disease was found mostly in plantations with planting materials and silvicultural treatments. Trees poor land preparation, where stumps and wood debris suffering from root disease were identified based on had been left on the ground to decay. The disease was observation of both above- and below-ground symp- also frequently observed in plantations with a pre- toms. The root system of suspected diseased trees vious history of root disease. Usually, untreated was excavated to examine for potential pathogens. disease centres were also associated with disease Pathogens were isolated from root samples in an incidence and it was considered likely that the path- attempt to identify the organisms causing the root-rot ogen in the new plantations originated from the dis- disease. Morphological studies were conducted both eased roots of previous crops. Root disease was low macroscopically and microscopically and fungal fea- or absent in plantations that has received good land tures were described according to Stalpers (1978). preparation by removing most of the stumps and Pathogenicity tests were conducted on 6-month-old wood debris (Table 1). Similar observations have seedlings using the method described by Mohd Farid been made by Van der Pas and Hood (1983) in Pinus et al. (2001). These tests were conducted under radiata plantations in New Zealand. natural field conditions using selected isolates of Rig- Overall, the majority of the plantations surveyed idoporus lignosus (Batu Anam, Johor), Rigidoporus were free from root disease. This appeared to be asso- vinctus (Pelong, Terengganu) and Phellinus noxius ciated with good land preparation and no previous (Lendu, Malacca and Tebuk Pulai, Selangor). history of root disease. Stumps and wood debris were mechanically removed and burned in the majority of Results and discussion these plantations. However, while root disease was not evident, symptoms of poor tree health, such as Disease survey stunted growth, small canopy size, small stems and leaves, multiple branching, and sparse and pale The surveys revealed that disease incidence was foliage, were common. The majority of teak and variable depending on tree species, land-manage- sentang growers considered poor tree health to have a ment practices and location. Root disease was found significant impact on production, particularly during in only 10 of 34 plantations surveyed (Table 1). Two the early phase of plantation establishment (Mohd main diseases were found: white root and brown root Farid et al. 2005). The condition is thought to be due disease. White root rot was more dominant on sen- to lateritic and/or compacted soil, which impedes the tang, while brown root disease was present in both growth of trees. Below ground, cracking of the root teak and sentang plantations. White root disease was surface and a lack of feeder roots were common recorded in monocultures of sentang at Sik, Kedah external signs of affected trees. The main anchoring and in sentang inter-planted with rubber trees in Batu root, however, was consistently found to be alive and Anam, Segamat, Ca’ah and Labis in Johor. Disease strong enough to support the tree. infection on sentang was recorded from as early as 1 year after planting. Similar results were gained by Root disease Maziah et al. (2001) with root-rot disease identified as a major threat to sentang plantations in the early Macroscopic and microscopic identification of stages of establishment (1–3 years old) and to rubber fruitbodies, infected roots and isolates, as well as trees 1–4 years old (Tan and Ismail 1991). pathogenicity tests, led to the identification of white Brown root disease was observed in teak planta- root disease and brown root disease caused by tions in Sabak Bernam, Selangor and Kuala Kangsar R. lignosus and P. noxius, respectively. The fungi in Perak as well as on sentang in Sik, Kedah and were the causal organisms as proven in the patho- Lendu, Malacca. Maziah and Lee (1999) have previ- genicity tests.

61 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. Land preparation (Yes/No) history of root disease root rubber) Yes Poor 6 rubber) No Good 7 × × Planting technique Previous Monoculture Yes Poor 3 MonocultureMonocultureMonoculture Yes Yes Yes Poor Poor Good 10 10 1 Mix (sentang Monoculture No Good 1 e d d d d Nil Monoculture No Good 8 P. noxius R. vinctus Phellinus noxius P. noxius NilNilNil Monoculture Monoculture Monoculture No No No Good Good Good 8 10 9 P. noxius Nil Monoculture No Good 4 Nil Monoculture No Good 18 NilRigidoporus lignosus Monoculture No Good 18 Pathogen Land management Age a b a b c Perlis Changlon, Kedah Changlon, Kedah Host, location, pathogen and land management of forest plantations in Peninsular Malaysia Sentang Km5. Labis Muar, Johor Nil Mix (sentang Sentang Lendu, Malacca Sentang Pelong, Terengganu TeakReserve, Forest Bintang Bukit 99. 1B/ & 99 1A/ Kompt. TeakTeak TeakTeak Kaki Bukit, Perlis Kaki Bukit, Perlis Timah Tasuh, Perlis Chuping, Perlis Nil Monoculture No Good 8 Host Site and state TeakTeakTeakTeak Sabak Bernam, Selangor Sabak Bernam, Selangor Perak River Estate, Kuala Kangsar, Kaki Bukit, Perlis SentangSentang Labis – Muar, Johor Ca’ah- Muar, Johor Nil Nil Monoculture Monoculture No No Good Good 5 5 TeakTeak Bukit Bintang Forest Reserve, Perlis Kompt. 22 & 23. Bukit Perangin Forest Reserve, Nil Monoculture No Good 16 Teak Kompt. 1, 2, 3 & 5 Bukit Perangin Forest Reserve, TeakTeakSentang Bukit Enggang Forest Reserve, Changlon, Perlis Tampin, Negeri Sembilan. Batu Anam, Segamat, Johor. Nil Monoculture Nil No Monoculture Good 21 No Good 7 Table 1.

62 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. Land preparation YesYes Poor Poor 7 5 (Yes/No) history of root disease root rubber rubber × × Planting technique Previous tree) tree) Monoculture Yes PoorMix (sentang Mix (sentang 7 e e e d NilNilNil Monoculture Monoculture Monoculture No No No Good Good Good 8 8 6 R. lignosus P. noxius R. lignosus R. lignosus Pathogen Land management Age Felda Titi, Negeri SembilanFelda Jengka, Pahang Unidentified Nil Monoculture Yes Monoculture Poor No 3 Good 1 (cont’d) Host, location, pathogen and land management of forest plantations in Peninsular Malaysia Distinguish between plantations located in the same district Brown root disease White root disease Sentang Khaya ivorensis Khaya ivorensis Ca’ah, Johor Host Site and state a,b,c SentangSentangSentangSentang KM 37.5, Labis- Yong Peng, Johor Km 0.5, Segamat-Muar, JohorSentang Bukit Hari, FRIM Sik, Kedah Kaki bukit, Perlis Nil Nil Monoculture Nild Monoculturee No Monoculture No Good Good 8 No 7 Good 5 SentangSentangSentangSentang Jementah, Johor Sentang Ulu Tiram, Johor Sentang Teluk Intan, Perak Bt 9, Jeniang, Kedah Sg. Chinoh Estate, Trolak Perak. Labis, Johor Nil Nil Nil Monoculture Monoculture Monoculture No No No Good Good Poor 5 6 6 Table 1.

63 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

These pathogens have wide host ranges and are scattered or solitary. In mixed plantations, dead trees responsible for most root and butt rot diseases in trop- were more clustered. ical rainforests (Lee 1997). Both R. lignosus and Brown root disease was found in both teak and P. noxius are economically important pathogens as sentang plantations. On teak, the disease caused basal they are responsible for causing more losses than all root rot (BRR) at Sabak Bernam, Selangor and root other pests and diseases combined, especially in rot at Kuala Kangsar, Perak. Its diagnosis was based rubber plantations (Fox 1977a; Johnston 1989). Rig- on rotting symptoms advancing up the root collar. At idoporus lignosus is economically important on present, BRR has been found only on teak at or above rubber and timber, especially in Indonesia, Malaysia, 10 years of age where it is planted on marine clay soil Sri Lanka and the Ivory Coast (Liyanage 1997a; known locally as Bernam series. Patches of dead Semangun 2000). It is the most destructive disease of trees were also an indicator of BRR. Below ground, rubber (Fox 1977b; Nandris et al. 1987a; Guyot and BRR was identified by the presence of a rough sheet Flori 2002) and the only disease that directly kills the of brown mycelial crust on the root surface. Soil par- tree irrespective of age and vigour (Tan and Ismail ticles, mainly of sand, frequently adhered to the crust. 1991). Phellinus noxius is also a destructive fungal In more advanced stages, a honeycomb structure of pathogen. It has been recorded on rubber, cocoa, tea, golden brown hyphae was formed in the rotted wood. fruit trees and forest trees (Pegler and Waterston These features have been observed and described in 1968; Singh 1973; Wood and Lass 1985; Nandris et detail by various other investigators (Anon. 1974; al. 1987a,b; Chang 1995; Ann et al. 2002). Nandris et al. 1983, 1987b; Ann et al. 1999). Two In the field, the above-ground symptoms associ- plantations of sentang located in Sik, Kedah and ated with each disease were similar; below-ground Lendu, Malacca were infected by brown root disease symptoms varied. Above ground, the symptoms were that killed both solitary trees and patches of trees irre- wilting, yellowing of leaves, loss of leaf lustre, bark spective of their health status. shrinkage, large canopy gaps, defoliation and die- One incidence of root disease was observed in back. The presence of these symptoms usually indi- K. ivorensis at Felda Titi, Negeri Sembilan caused by cated that the trees were beyond the point of an unidentified fungus. Both below-ground and recovery, as the fast progress of infection leads to above-ground observations revealed that the disease rapid death (Ismail and Azaldin 1985). Spread of symptoms were similar to brown root disease caused both diseases to adjacent healthy trees was primarily by P. noxius, with some small variations. The surface through root contact. This is the most common mode of the infected tree root was often covered with a of disease-spread in plantation-grown rubber (Anon. brown mycelial crust with adhering soil particles. 1974; Holliday 1980; Nandris et al. 1983, 1987a; The crust was usually whitish in colour when young, Rajalakshmy and Jayarathnam 2000), teak (Tewari becoming brownish over time. Further study is 1992), Acacia mangium (Lee 1997; Maziah 2002; needed to identify the unknown pathogen. Ito, unpublished data) and Douglas-fir (Wallis and Reynolds 1965). Frequently, the source was infected old stumps or wood debris remaining in the soil or Control and management of root standing diseased trees. In rubber plantations, the disease source of inoculum for root disease infection is mainly from infected rubber trees, stumps or forest At present, the incidence of root disease in forest trees (Rajalakshmy and Jayarathnam 2000). This is plantations in Peninsular Malaysia is relatively low similar to root rot observed on A. mangium by Old et compared with that in rubber tree and oil-palm plan- al. (2000). tations. This destructive disease has the potential, White rhizomorphs on the root surface were the however, to be a major threat to the timber plantation main indicator in identifying R. lignosus in the field. industry in the future. Experience gained in the con- Their presence means that the whole root system has tainment and control of root disease in rubber planta- already been exposed to the disease (Wheeler 1974). tions could provide useful strategies to prevent the The disease survey showed that white root disease spread of the disease in forest plantations. occurred in both sentang monocultures and sentang In rubber plantations, root diseases are managed by inter-planted with rubber. In monocultures, disease cultural practices, especially through land clearing incidence was relatively low and diseased trees were (Old et al. 2000). Therefore, during the conversion of

64 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. land for the establishment of timber tree plantations, Based on the experience of rubber-tree growers in stumps and woody debris should be removed and the control of white root rot disease caused by destroyed. The aim of this activity is to reduce the R. lignosus, good land clearing during site prepara- source of potential inoculum in the soil. This tion, the construction of isolation trenches and the approach could minimise the incidence of the disease application of fungicides are recommended as valu- in the plantations (Fox 1977b; Liyanage and Peries able tools to help manage root rot disease in forest 1982). In rubber plantations, the construction of iso- tree plantations. lation trenches around trees identified as diseased is another recommended control strategy. This is the References most common and popular method of root disease control to prevent disease spread to adjacent healthy Ann, P.J., Lee, H.L. and Huang, T.C. 1999. Brown root rot of trees through root contact (Maziah and Lee 1999). 10 species of fruit trees caused by Phellinus noxius in However, the method is costly and labour intensive, Taiwan. Plant Disease, 83, 746–750. especially when large areas or many patches of Ann, P.J., Chang, T.T. and Ko, W.H. 2002. Phellinus noxius disease occur in the plantation. brown rot of fruit and ornamental trees in Taiwan. Plant The most common practice of rubber growers to Disease, 86, 820– 826. combat white root disease is the application of fungi- Anon. 1974. Root diseases of Hevea. Planter’s Bulletin of cides by means of soil drenching. Several fungicides, the Rubber Research Institute Malaysia, 133, 109–200. such as hexaconazole, tridemorph, propiconazole, Browne, F.G. 1968. Pest and disease of forest plantation trees. An annotated list of the principal species occurring tridemefon, cyproconazole and penconazole, have in the British Commonwealth. Oxford, Clarendon Press, shown promise in the control of disease caused by 1330p. R. lignosus. The efficacy of the fungicide treatment, Chang, T.T. 1995. A selective medium for Phellinus noxius. however, reduces with increasing levels of infection European Journal of Forest Pathology, 25, 185–190. (Ismail and Shamsuri 1998). Fungicides should Fox, R.A. 1977a. The role of biological eradication in root therefore be applied only on newly infected trees or disease control in replantings of Hevea brasiliensis. In: trees at mild infection levels. Baker, K.F. and Snyder, W.C., ed., Ecology of soil-borne plant pathogens. Berkeley, University of California Press, 348–362. Conclusion — 1977b. The impact of ecological, cultural and biological factors on the strategy and costs of controlling root Two major root diseases of forest plantation species, diseases in tropical plantation crops as exemplified by white root (R. lignosus) and brown root (P. noxius), Hevea brasiliensis. Journal of Rubber Research Institute were found during the surveys. Sentang was suscep- Sri Lanka, 54, 329–362. tible to white root disease, especially when inter- Guyot, J. and Flori, A. 2002. Comparative study for planted with rubber trees. Brown root disease, in detecting Rigidoporus lignosus on rubber trees. Crop comparison, was found in both teak and sentang plan- Protection, 21, 461–466. tations. A root disease caused by an unidentified Holliday, P. 1980. Fungus diseases of tropical crops. fungus was found on khaya at Felda Titi in Negeri Cambridge University Press, 607p. Sembilan. This pathogen killed its host aggressively Ismail, H. and Azaldin, M.Y. 1985. Interaction of sulphur without showing early symptoms. Trees infected by with soil pH and root diseases of rubber. Journal Rubber this disease were almost indistinguishable from Research Institute Malaysia, 33, 59–69. healthy trees, with symptoms discernible only during Ismail, H. and Shamsuri M.H. 1998. Current status of root the advanced stages of infection. disease of rubber. Paper presented at CABI workshop on Ganoderma diseases, 5–8 October 1998. Serdang, The survey also revealed that infection started MARDI Training Center. mainly from infected roots of trees, stumps or root Johnston, A. 1989. Diseases and pests. In: Webster, C.C. and remnants remaining in the soil, and that spread to Baulkwill, W.J., ed., Rubber. New York, Longman adjacent trees was through root contact. Good land Scientific and Technical, 415–458. preparation by removing all woody debris and Lee, S.S. 1993. Diseases. In: Kamis, A. and Taylor, D., ed., stumps in the soil before plantation establishment Acacia mangium growing and utilisation, 203–223. was therefore considered vital to reduce disease inci- Bangkok, Winrock International and FAO, MPTS dence. Monograph Series No. 3, 203–223.

65 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

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66 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

The biological control of Ganoderma root rot by Trichoderma

S.M. Widyastuti1

Abstract This paper describes the ability of Trichoderma spp. to act as agents for biological of Ganoderma. Certain species of Ganoderma are potential root-rot pathogens and are capable of causing serious damage to many types of plantation-tree species in Indonesia. Biological control of plant pathogens aims to decrease dependence on chemical treatments which may cause environmental pollution and the development of resistant strains. Filamentous fungi such Trichoderma that are mycoparasites of plant pathogens have potential for the biocontrol of plant disease. Species of Trichoderma are one of the most widely tested agents. Although the mechanism of mycoparasitism is not fully understood, expression of extracellular cell-wall degrading enzymes is assumed to be involved in this process, including the action of chitinolytic and glucanolytic enzymes. As reported for other chitinolytic systems, the endochitinase (EC 3.2.1.14) is among the most effective for both antifungal and lytic activities in comparison with other types of chitinolytic enzymes. Recently, 32-kDa endochitinolytic enzymes have been purified from Trichoderma reesei and characterised. We have tested the antagonistic ability of Trichoderma isolates against some plant pathogenic fungi, such as Ganoderma spp., Rigidoporus microporus, Rhizoctonia spp., Fusarium sp., and Sclerotium rolfsii. Results show that Trichoderma spp. can suppress the development of fungal pathogens in vitro and in glasshouse experiments.

Intensive forestry is based on growing one or few tree soil-borne disease are limited, and measures must be species over large areas. These plantation ecosys- developed to avoid the start of an epidemic by pre- tems are ecologically unbalanced and favour epi- venting inoculum build-up. demics of pathogens or pests that interfere with the In agriculture, and to some extent forestry, the production of a healthy, valuable tree crop. Preven- chemical treatment of pests has been replaced or tion of such epidemics in forest plantations cannot be reduced through the use of biologically based fungi- achieved through the use of chemical fungicides cides. A broad definition of biological control is the since it would not be cost-effective or environmen- reduction of the amount of inoculum or disease-pro- tally sustainable. Consumers are increasingly con- ducing activity of a pathogen accomplished by or cerned about the chemical pollution of the through one or more organisms other than humans environment, and pathogens could become resistant (Cook and Baker 1983). This definition includes the to available chemicals if these are used indiscrimi- use of less-virulent variants of the pathogen, more nately. The options currently available to manage resistant cultivars of the host, and microbial antagonists that interfere with the survival or disease-producing activities of the pathogen. This paper 1 Faculty of Forestry, Gadjah Mada University, discusses the development of a biological control for Bulaksumur, Yogyakarta 55281, Indonesia. Ganoderma rot root disease using antagonistic fungi Email: . of the genus Trichoderma.

67 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

What are Ganoderma? possible by chemical fungicides. Trichoderma can be used to attack established pathogens as well as pre- The Ganodermataceae are cosmopolitan basidiomyc- venting the establishment of disease. etes which cause root rot of many temperate and tropical hardwoods by decomposing lignin as well as cellulose and related polysaccharides (Hepting 1971; Antagonistic mechanisms of Blanchette 1984; Adaskaveg and Ogawa 1990; Trichoderma Adaskaveg et al. 1991, 1993). Successive replanting of monocultures such as Acacia mangium Willd. in In addition to colonising roots, Trichoderma spp. Southeast Asia can be rapidly exploited by soil-borne attack, parasitise and otherwise obtain nutrition from fungi such as Ganoderma, and this particular other fungi. Since Trichoderma spp. grow and prolif- problem will become more serious as more areas erate when there are abundant healthy roots, they move into second or even third-rotation planting. have evolved numerous mechanisms to enable them Environmental considerations also mean that native to attack other fungi and to enhance plant and root forest areas can no longer be exploited, making growth. further replanting of these plantation forests inevi- A list of recently described mechanisms follows: table. It is thus essential to develop appropriate, inte- • mycoparasitism — in which one fungus derives its grated management systems for root rot diseases. nutrition from another without any benefit in Ganoderma spp. have been found worldwide on a return. The interaction can be where the parasite is range of broad-leaved hosts (Phillips and Burdekin biotrophic or necrotrophic as is the case for 1989). Butt rot and root rot symptoms of Ganoderma Trichoderma. spp. have been recognised on planted Acacia in • antibiosis — an association between two northern Australia (C. Mohammed, pers. comm.), in organisms that is detrimental to the vital activities a provenance trial in Peninsular Malaysia, in northern of one of them and, in fungi, is usually mediated by Sumatra (Lee 1996) and elsewhere in Indonesia toxic metabolites produced by one organism (Irianto et al. 2006). Root-rot disease caused by Gan- • competition for nutrients or space — the active oderma spp. has been reported as the most serious requirement for resources in excess of those disease of A. mangium in West Bengal, India immediately available to two or more organisms. (Sharma and Florence 1996). The production of toxic metabolites is known to be affected by the nutrient status of the growth What are Trichoderma? medium (Ghisalberti and Sivasithamparan 1991; Howell and Stipanovic 1995). Trichoderma spp. are fungi that are often dominant • tolerance to stress through enhanced root and plant components of the soil microflora in widely varying development habitats. This may be attributable to their diverse • solubilisation and sequestration of inorganic metabolic capability and their aggressively competi- nutrients tive nature. Strains of Trichoderma are rarely associated with disease in living plants. • induced resistance The high degree of ecological adaptability shown • inactivation of the pathogen’s enzymes. by strains within the genus, coupled with their ame- Trichoderma strains produce a variety of volatile nability to cultivation on inexpensive substrates, and non-volatile toxic metabolites. Of these, some make Trichoderma isolates attractive candidates for are considered to be antibiotics as they can inhibit the a variety of biological control applications (Hjeljord growth of other microorganisms without physical and Tronsmo 1998). As fast-growing saprophytes, contact between the fungi. The best known of the Trichoderma can compete ecologically over the long antifungal metabolites produced by isolates of this term as well as at the time of application and are able genus is the coconut-scented 6-n-pentyl-2H-pyran-2- to colonise potential infection courts, such as one (PPT) (Claydon et al. 1987). In addition, many growing roots, wounds, or senescent tissue, as they strains of Trichoderma are able to produce extracel- become available. As living organisms, biological lular cell-wall-degrading enzymes which are also control agents can act as aggressive mycoparasites capable of killing at a distance. These, however, are and adapt to changes in their habitat in a manner not traditionally included in the concept of mycopara-

68 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. sitism, due to their integral role in direct physical The type of antagonism that appeared the most effec- interactions. tive in inhibiting the root-rot pathogens tested was mycoparasitism (when the Trichoderma grew within Trichoderma as biological control the pathogen’s colony (Figure 1a)). Antagonism by competition occurred when both the pathogen and agents for agents of root-rot disease Trichoderma grew but the growth of Trichoderma including Ganoderma limits the full access of the pathogen to the substrate (Figure 1b). No one colony was able to dominate the In order to test the potential antagonistic effects of Tri- substrate once hyphal contact was established choderma spp. against various root-rot pathogens (Figure 1b). Interactions typical of antibiosis were causing problems in Indonesia, we prepared cultures by observed, i.e the Trichoderma isolate is producing placing agar plugs containing the mycelia of the two toxic compounds and the colony of the root-rot path- antagonists (Trichoderma and root-rot pathogen) on ogen is severely restricted and clearly contained by opposite sides of the agar plate. The plant pathogenic the Trichoderma (see Figure 1c in comparison to 1b). fungi tested against Trichoderma spp. isolates were In another experiment, three Trichoderma isolates, Ganoderma spp. (Widyastuti and Sumardi 1998; Wid- previously shown to have high antagonistic ability yastuti et al. 1998a, 1999), Rigidoporus microporus (T. koningii isolate T1, T. reesei isolate T13 and (Widyastuti et al. 1998b), Rhizoctonia spp. (S.M. Wid- T. harzianum isolate T27; Figure 2) were tested yastuti, Harjono, Sumardi and N.S. Lestari, unpub- against isolates of Ganoderma collected from dif- lished data; S.M. Widyastuti, Sumardi, Harjono and E. ferent tree species. It was significant, however, that Windyarini, unpublished data), Fusarium sp. (S.M. these Trichoderma isolates of known high antago- Widyastuti, Sumardi and Y. Mitikauji, unpublished nistic ability actually varied greatly in their level of data) and Sclerotium rolfsii (Widyastuti et al. 2003). antagonism towards the different Ganoderma iso- Control plates were inoculated with only one of the lates. Widyastuti et al. (1999) showed in paired cul- antagonists but on both sides of each individual plate in tures that a Trichoderma isolate which was highly order to simulate growth conditions as comparable as effective against one isolate of a pathogen could have possible to those in the test plates. Replicates com- minimal effect on other isolates of this pathogen prised of a minimum of three plates per combination of (Table 1). This might be related to the high path- each pair of fungi. Interaction zones, i.e. the areas of ogen–Trichoderma specificity of antagonistic mech- contact and subsequent overlap of hyphae of Tri- anisms due to antibiosis (Howell and Stipanovic choderma spp. and pathogenic fungi, were observed at 1995) and cell-wall-degrading enzymes (Haran et al. various magnifications in situ by light microscopy. 1996). This specificity theory is most probable since Among the 120 isolates of Trichoderma spp. tested the Ganoderma isolates could have been different against the pathogenic fungi we recorded examples isolates of the same Ganoderma species or isolates of of the basic types of antagonistic behaviour — anti- different Ganoderma species and their identity will biosis, mycoparasitism, and competition (Figure 1). be tested in future work.

Figure 1. Three basic interaction mechanisms between Trichoderma and pathogenic fungal isolates: (a) mycoparasitism, (b) competition, (c) antibiosis. Key: T = Trichoderma, P = Pathogen.

69 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Microscopic observation of koningii. The above responses observed (growing antagonistic interactions between parallel to the pathogen, coiling, appresoria, hooks) are considered to be examples of mycoparasitic Ganoderma philippi and Trichoderma activity and are similar to those described by Elad et al. (1983), who observed the mycoparasitic activity Hyphal interactions were observed under a light microscope with hyphae that had been stained in 10% of T. harzianum against S. rolfsii and R. solani using (v/v) lactophenol blue for 5 minutes. Observations of scanning electron and fluorescence microscopy. the zone of the confrontation between Trichoderma Trichoderma reesei was the most effective mycopar- and Ganoderma philippi isolates indicated a range in asite in interactions with Ganoderma isolates, followed the types of hyphal interactions. The diameter and the by T. koningii and T. harzianum (Widyastuti et al. intensity of staining of Ganoderma and Trichoderma 2003; Figures 2–3 in this paper). In interactions with fungal hyphae were different, so they were easily dis- pathogenic fungi other than Ganoderma, T. koningii tinguished from each other (Figure 3a–c). All Tri- has displayed a typical antibiosis inhibition pattern of choderma isolates were frequently observed to grow confinement (e.g. Figure 1c), whereas in interactions parallel to the pathogen. Hyphae of T. reesei and with Ganoderma the T. koningii isolate T1 demon- T. harzianum often coiled around the pathogen strated limited mycoparasite properties (with hyphae (Figure 3a,b), but no coiling was shown by T. kon- growing parallel to the pathogen). This isolate T1 can ingii (Figure 3c). Trichoderma reesei and T. har- most probably exhibit both main types of fungal inhi- zianum both produced appresoria at the tips of short bition, mycoparasitism and antibiosis depending on the branches (Figure 3d) or formed a hook-like structure particular species of fungal pathogens with which it is (Figure 3e). No such structures were produced by T. interacting (Widyastuti et al. 2003).

Table 1. Growth inhibition of Ganoderma spp. from eight different tree species in co-culture with three Trichoderma spp. isolates

Isolate Tree species Mean growth inhibition (%) number T. koningii (T1) T. reesei (T27) T. harzianum (T13) 13 Paraserianthes falcataria 62.8 100.0 56.4 24 Leucaena leucochepala 99.5 100.0 89.4 2 Acacia mangium 94.3 96.5 79.8 9 A. auriculiformis 96.4 98.4 86.6 19 Cassia siamea 98.6 98.3 90.6 10 Delonix regia 97.0 98.2 83.0 4 A. mangium 98.8 98.2 84.6 17 Dalbergia latifolia 98.5 95.7 71.6

Figure 2. Antagonistic ability of three Trichoderma isolates varied towards pathogenic fungi (Ganoderma sp.) grown in paired cultures on agar: (a) Ganoderma sp. vs T. Reesei; (b) Ganoderma sp. vs T. Koningii; (c) Ganoderma sp. vs T. harzianum. Key: P = Pathogen, T = Trichoderma

70 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Extra-cellular enzyme production in The colony development of G. philippii is clearly Trichoderma–Ganoderma philippi inhibited at concentrations of 90–200 mg/mL (Figure 4a). A concentration of 80 µg/mL shows only an interactions obscure inhibition zone. No inhibition zone is Extracellular cell-wall-degrading enzymes such as observed at concentrations of 40–70 µg/mL. chitinolytic and glucanolytic enzymes are believed to Microscopic observations revealed that morpho- be involved in mycoparasitism. Recently, 32-kDa logical changes in G. philippii hyphae were induced endochitinolytic enzyme was purified and character- by T. reesei 32-kDa endochitinase (Table 2). ised from T. reesei (Harjono et al. 2001; Harjono and Branching and segmentation of hyphal tips occurred Widyastuti 2001a) and tested for its ability to inhibit at 60 µg/mL of endochitinase application (Table 2 colony and hyphal growth of G. philippii isolate T13 and Figure 4b,c). At this concentration, the enzyme (Harjono and Widyastuti 2001b). also caused swollen hyphal tips (Figure 4d).

Figure 3. Light microscopy of Trichoderma hyphae interacting with that of Ganoderma: (a and b) Condensed coiling of T. reesei and T. harzianum, respectively, around a hypha of Ganoderma sp. (c) T. koningii could grow parallel to hypha of Ganoderma sp. but no coiling was observed. (d) Appresorium-like structures, formed by T. reesei, also found in T. harzianum but not in T. koningii. (e) Hook-like structure found in T. reesei and T. harzianum which serve as an attachment to the host mycelium. (Bar represents 10 µm).

71 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Hyphal necrotic lesions were observed at concentra- T. harzianum and T. koningii delayed the initiation of tions 80, 90, 100 and 200 µg/mL (Table 2). The latter symptoms and decreased disease incidence by 73%, 32-kDa endochitinase concentrations caused perma- 67% and 47%, respectively, when applied 4 days nent hyphal necrosis of G. philippii (Figure 4e). Hyphal before disease inoculation. Disease reduction by Tri- lysis by the enzyme was observed at a concentration of choderma declined drastically when Trichoderma was 90 µg/mL and the mean percentage increased dramati- applied 4 days after inoculation with S. rolfsii. Adapta- cally at a concentration of 200 µg/mL (Table 2). At this tion and establishment of Trichoderma before path- high concentration, cell bursting and protoplast release ogen inoculation probably increases antagonistic was observed not only on hyphal tips (Figure 4g), but capability. Our results agree with findings of Widyas- also on mature hyphae (Figure 4h). Enzyme applica- tuti et al. (2002) that formulated Trichoderma needs a tion at low concentrations (80 µg/mL) failed to sup- period of adaptation in order to actively inhibit the press the growth of hyphae (Figure 4f). growth of pathogenic fungi.

Efficacy of formulated Trichoderma Table 3. Control by three isolates of Trichoderma as a biological control agent tested formulated in alginate beads of damping- off caused by Sclerotium rolfsii in against Sclerotium rolfsii greenhouse-grown pine seedlings

Greenhouse experiments were used to evaluate the effi- Trichoderma isolates Disease reduction accord- cacy of isolates T13 (T. reesei), T27 (T. harzianum) and ing to S. rolfsii inocula- a T1 (T. koningii) as biological control agents in a formu- tion lation of alginate beads (Widyastuti et al. 2003, 4 days At the 4 days 2006a,b,c), against a common damping-off pathogen, before same after Sclerotium rolfsii. The levels of biological control time achieved for the damping-off by S. rolfsii on pine seedlings by applying Trichoderma spp. to soil are recorded T. koningii isolate T1 46.7b 43.3ab 3.3b in Table 3. The application of formulated Trichoderma T. reesei isolate T13 73.3a 50.0a 23.3a T. harzianum isolate T 66.7a 56.7a 6.7b into potting soil 4 days before the pathogen S. rolfsii 27 was inoculated gave the best disease suppression, with a Data in each column which are followed by a common letter are not statistically different (p=0.05). Percentage disease the Trichoderma delaying the initiation of symptoms reduction was calculated by counting the number of dead or and reducing disease incidence. Trichoderma reesei, dying plants in 15-day-old pine seedlings.

Table 2. Microscopic observations of the morphological anomalies in G. philippii hyphae caused by 32-kDa Trichoderma reesei endochitinasea

Endochitinase Appearance of abnormal hyphaeb concentration Segmented Branched Swollen Necrotic Lysed (µg/mL) 0 – – – – – 40 – – – – – 50 – – – – – 60 1.7 0.8 1.8 – – 70 1.7 2.5 0.8 – – 80 4.2 2.5 2.5 5.0 – 90 6.7 5.0 2.5 8.3 1.7 100 10.8 5.8 7.5 11.7 9.2 200 7.5 8.3 6.7 15.8 14.2 a Data obtained 12 hours after enzyme application. b Percentage of 120 hyphae observed under microscope (mean of two replications). – Indicates that at these concentrations the endochitinase did not induce a morphological response in Ganoderma hyphae.

72 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Figure 4. Effects of the 32 k-Da T. reesei endochitinase on hyphae of G. philippii: (a) Growth inhibition of G. philippii in response to various endochitinase concentrations (from top, in clockwise direction, 50, 100, 200, 0 µg/mL); (b, c, d) Branching, segmentation and swollen hyphal tip of G. philippii in the presence of endochitinase at 60 µg/mL and above; (e) G. philippii hyphae necrotic at hyphal tip grown in the presence of endochitinase higher than 70 µg/mL; (f) Hyphae of G. philippii after enzyme exposure of 80 µg/mL showing recovery after swelling and continued normal hyphal growth; (g) Cell bursting and protoplast release of hyphal tips in the presence of endochitinase at 90 to 200 µg/mL; (h) Cell bursting and protoplast release of mature hyphae. (Bars = 30 µm)

Conclusion of Ganoderma root rot without using chemical fungicides and the ensuing dangers of environmental pol- This paper provides an overview of various aspects lution and the development of fungicide resistance. of the antagonism of Trichoderma isolates that are relevant to their use as biological control agents. We Acknowledgments are now able to screen Trichoderma isolates for traits relevant to antagonism, and select isolates according I thank Harjono Djoyobisono and Ananto Triyogo to their potential for biological control. Applied in for their help in preparing this manuscript. appropriate biologically based formulations and delivery systems and at the right stage of the disease cycle, specific Trichoderma isolates selected for high References levels of Ganoderma- suppressive activity warrant Adaskaveg, J.E. and Ogawa, J.M. 1990. Wood decay further investigation. Such Trichoderma isolates pathology of fruit and nut trees in California. Plant offer the potential for consistent and effective control Disease, 74, 341–352.

73 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

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Asian Journal of Sustainable Agriculture, endokhitinase dari jamur mikoparasit Trichoderma 1(2), 1–8. reesei (Optimization of endochitinase production from Widyastuti, S.M., Sumardi and Harjono 1999. Potensi mycoparasitic fungi Trichoderma reesei). Indonesian antagonistik tiga Trichoderma spp terhadap delapan Journal of Plant Protection, 7, 52–55. penyakit akar tanaman kehutanan (Antagonistic potential — 2001b. Antifungal activity of purified endochitinase of three Trichoderma spp. to suppress eight root-rot produced by biocontrol agent Trichoderma reesei against diseases of forest plant). Forestry Bulletin, 4, 2–10. Ganoderma philippii. Pakistan Journal of Biological Widyastuti, S.M., Sumardi and Hidayati, N. 1998b. Science, 4, 1232–1234. Kemampaun Trichoderma spp. untuk pengendalian Harjono, Widyastuti S.M. and Margino, S. 2001. Pemurnian hayati jamur akar putih pada Acacia mangium secara in dan karakterisasi enzim endokitinase dari agen vitro. (In vitro efficacy of Trichoderma spp. to control pengendali hayati Trichoderma reesei (Purification and white root-rot isolated from Acacia mangium). Forestry characterization endochitinase enzyme from biocontrol Bulletin, 36, 24–38. agent Trichoderma reesei). Indonesian Journal of Plant Widyastuti, S. M., Sumardi, Irfa’I and H. H. Nurjanto. 2002. Protection, 7, 114–120. Aktivitas penghambatan Trichoderma spp. formulasi Hepting, G.H. 1971. Disease and forest and shade trees of terhadap jamur patogen tular tanah secara in vitro. (In the United States. US Department of Agriculture, vitro inhibition activity of formulated Trichoderma spp. Agricultural Handbook, 386, 1–658. against soil-borne pathogenic fungi). Indonesian Journal Hjeljord, L. and Tronsmo, A. 1998. Trichoderma and of Plant Protection 8, 27–34. Gliocladium in biological control: an overview. In: Widyastuti, S.M., Sumardi, A., Sulthoni and Harjono. Harman, G.E. and Kubicek, C.P., ed., Trichoderma and 1998a. Pengendalian hayati penyakit akar merah pada Gliocladium: enzymes, biological control and akasia dengan Trichoderma (Biological control of red– commercial applications (volume 2). London, UK, root rot disease of acacia using Trichoderma). Indonesian Taylor and Francis Ltd, 393p. Journal of Plant Protection, 4, 65–72. Howell, C.R. and Stipanovic, R.D. 1995. Mechanisms in the biocontrol of Rhizoctonia solani induced cotton seedling

74 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Minimising disease incidence in Acacia in the nursery

Budi Tjahjono1

Abstract Disease management in the nursery requires detailed knowledge of the potential pathogens and their interactions with the seedlings and their environment. The behaviour of the pathogen must also be considered in the wider context of the nursery operation. The conditions that favour a range of foliar, stem and root diseases of Acacia seedlings in the nursery are described. Integral control strategies are discussed in general, and in detail for Xanthomonas sp., the common bacterial leaf blight of Acacia crassicarpa.

Reforestation of the industrial plantation forest (or to fungicides, all contribute to the development of HTI) in Indonesia should be undertaken by planting control strategies. seedlings immediately after an area has been cut Disease management must be considered in the over. Implementing this recommendation, however, context of the wider nursery operation: container requires the timely supply of seedlings and often management, shading, irrigation, fertiliser applica- creates a demand for high rates of production from tion, pest control and sanitation. Most diseases in plantation nurseries. To attain this goal, uniform, vig- nurseries can be adequately managed by the applica- orous, high-quality seedlings must be produced and tion of good cultural practices. Poor practices, such disease incidence and losses kept to a minimum. as incorrect sowing depth, high seedling density and Seedling losses in the nursery directly affect rates of inappropriate levels of shade, can increase the inci- plantation establishment, as well as increasing main- dence and severity of diseases. tenance costs during the rotation. Crowding of seedlings encourages the development of some diseases that are much reduced or dis- Nursery management appear after the stock has been transplanted in the field. Whether the seedlings are grown under shade It is important to identify the actual and potential net, plastic protection or the open sky also affects the disease problems in the nursery in order to imple- occurrence of nursery diseases. Excessive shade, ment an effective disease-management strategy. resulting in low light intensity, tends to lower soil Detailed knowledge of the pathogen and its interac- temperature and raise the water potential, favouring tion with the seedling and environment is vital. Infor- the development of damping-off disease. Con- mation on the origin of the pathogen, how it is spread, versely, seedbeds under high light intensity tend to the severity of its impact, what environmental and have higher soil temperature and lower soil-water host factors favour its development, and its response potential, conditions that favour the development of seedling blight and shoot blight. 1 PT. Riau Andalan Pulp and Paper, PO Box 1089, The selection of growing medium is also important Pekanbaru 28000, Indonesia. in nursery disease management. Seedbed soils and con- Email: . tainer mixes for acacia seedlings need to have good

75 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. structure, be free-draining and slightly acid (pH5–5.5). spot (Figure 1a), Phaeotrichoconis leaf spot (Figure Poor drainage causes waterlogging damage and can 1b) and bacterial leaf blight caused by Xanthomonas lead to the development of soil-borne diseases. An sp. Important leaf diseases on Acacia mangium ideal medium for container production of acacia seed- Willd. seedlings are phyllode rust disease caused by lings has an organic base to support the nursery produc- Atelocauda digitata (Figure 2), anthracnose disease tion phase (about 10–12 weeks), with sufficient added and tip necrosis caused by Colletotrichum sp. fertiliser to reduce follow-up fertiliser application to a minimum. Organic-based media also provide a recep- Stem and root diseases tive base should inoculations of mycorrhizae or biological control agents be required. Acacia seedling growth From germination through the first few weeks after is generally restricted in the nursery after 10 weeks emergence, the succulent radicle and hypocotyl through reduced irrigation and fertiliser. If seedlings tissues of Acacia seedlings are extremely susceptible are retained in the nursery for more than 12 weeks, to attack by damping-off fungi. Pythium, Rhizoctonia there is potential for severe levels of diseases to and Fusarium are fungi that commonly cause develop. damping-off. The exact cause of many root diseases is often difficult to determine without laboratory analysis. Pre-emergence damping-off is caused by fungi that rot seedlings before they emerge. Post-emergence damping-off is caused by fungi that infect and kill stem tissues at ground level after seedling emergence, resulting in seedling collapse. Because damping-off is significantly influenced by environmental conditions, the severity of this disease fluctuates from year to year. In general, conditions that reduce seedling growth and vigour predispose nursery stock to increased infection from damping-off fungi. Nitrogen fertilisers applied before or during the first few weeks after seedling emergence may also increase infection. Other diseases of seedlings include canker and dieback pathogens. These attack the stems and branches of nursery seedlings, cause sunken lesions and malformations, and often lead to seedling death.

Integrated disease management

Figure 1. Common leaf diseases in Acacia The basic approach to disease management in the crassicarpa: (a) Pestalotiopsis leaf spot; nursery should be to avoid disease rather than have to (b) Phaeotrichoconis leaf spot apply controls after a disease outbreak. Common ways to minimise disease in nurseries that raise Foliage diseases Acacia seedlings include: • use of high-quality seeds from genetic material For an infection to be established, fungi and bacteria that has disease resistance that cause foliage diseases usually require conditions • cultural control through growing conditions, of high moisture and free water in and around stems media and cultural practices and foliage for long periods. Foliage diseases are • proper irrigation and drainage therefore most prevalent in locations where frequent • good hygiene and quarantine rain or the continuous use of overhead irrigation • the application of chemical or biological control, allows these conditions to develop. including selective pesticides that kill or inactivate Common leaf diseases in Acacia crassicarpa the pathogen but cause relatively no harm to the A. Cunn. ex Benth. seedlings are Pestalotiopsis leaf seedlings.

76 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Figure 2. Phyllode rust caused by Atelocauda digitata on Acacia mangium seedlings

Figure 3. Common bacterial leaf blight in A. crassicarpa seedlings: (a) symptoms; (b) yellow colony of pathogenic bacteria Xanthomonas sp.; (c) leaf with bacterial ooze; (d) cross-section of infected leaf (400× magnification)

Several cultural practices can reduce or control lings. Watering should be done only as needed and the nursery diseases. Good-quality growing media and irrigation systems should be regularly inspected for full containers encourage the growth of vigorous leaks or blockages. Foliage diseases in particular can seedlings that are better able to resist stress and dis- be inhibited by reducing seedbed density in order to ease. Good hygiene and housekeeping, based on the lower humidity and increase air circulation around the nursery area being kept well-drained, clean and leaves. The removal and controlled burning of heavily orderly, reduce disease infection and loss of seed- infected and dead seedlings from seedbeds will

77 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. reduce the source of inoculum, as well the likelihood (Figure 3). Acacia mangium and Acacia auriculi- of subsequent infection of other nursery seedlings. formis A. Cunn. ex Benth. appear immune to this dis- Chemical or biological pesticides may be used to ease. Conditions that predispose seedlings to the protect seedlings from infection or eradicate a path- development of bacterial leaf blight include: ogen already present in the nursery. Spraying with a • stress due to excess or lack of water, lack or fungicide, for instance, protects the seedlings by imbalance in nutrition, unsuitable growing media coating the foliage with a chemical barrier that is and/or excess nitrogen fertiliser application toxic to foliar pathogens. Such chemical control • poor sanitation of media, water supplies and tube requires proper timing to coincide with periods of sterilisation potential infection. Soil fumigation before planting • intense rainfall or strong splash water from can control root-disease pathogens, but seldom will it irrigation completely eradicate them. • leaf scars/lesions as entry points for the bacteria. The concept of integrated disease management in The recommended steps for integrated control are: the nursery can be represented by the palm of the • use suitable growing media with adequate porosity hand and its five fingers. Each finger represents one but high water retention of the five components of disease control (see dot • water only when required and ensure fertiliser points above); the palm is the nursery manager. The applied is balanced palm is central to coordinating the five fingers. A • maintain good nursery hygiene with proper single approach to disease management in the drainage nursery seldom leads to disease control. • remove and burn dead and severely diseased plants • handle seedlings gently to reduce scars and lesions The integrated management of • sterilise (using steam if possible) equipment, common bacterial leaf blight — seedbed, containers a case study • apply bactericide: Agrept 20 WP and Kibox (copper oxychloride) Bacterial leaf blight of A. crassicarpa caused by Xan- • plant non-host plant material, i.e. species other thomonas sp. is a common problem in the nursery than A. crassicarpa.

78 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Developing a strategy for pruning and thinning Acacia mangium to increase wood value

Chris Beadle1

Abstract

Timber production from Acacia mangium plantations in Indonesia can potentially supply both domestic and export markets for furniture and other functional and appearance products. In plantations, there is potential for large and persistent branches to develop, and the threat of persistent dead branches and of heart rot. This has resulted in the need for pruning systems based on the removal of green branches from below, and thinning systems that ensure final-crop trees retain green branches until pruning is completed, as well as maintaining acceptable rates of growth of the retained trees. This paper outlines a pruning and thinning strategy for A. mangium that attempts to meet the criteria required for growing high-quality, clear-wood plantations. Form- pruning to encourage the development of good form ahead of lift-pruning is used in a system that results in about 300 stems/ha of trees pruned to 4.5 m in the final crop. It is concluded that heart rot is made worse by pruning when the plant material is susceptible and a sufficient source of fungi is present to invade pruning wounds.

Acacia mangium Willd. plantations represent a sig- Single stems are required for plantation silviculture nificant proportion of the wood supply to the pulp and and are essential for solid-wood production. A pruning paper industry in Indonesia (Rimbawanto 2002). The strategy therefore becomes necessary. species is fast growing and of medium density. When When managed for pulpwood, A. mangium planta- free from defects, the wood looks similar to teak tions are established at around 1000 stems/ha. Such (Gales 2002). The timber is also used in the domestic planting densities are too high for all the trees to be market in Indonesia for furniture and other functional managed for solid wood. A lower stocking at and appearance products, and exported for finger- planting is one option but experience from growing jointing in a developing overseas market (Hardiyanto other species suggests that starting with higher stock- 2006). With relatively short rotations of no more than ings ensures that there are sufficient potential final- 20 years to produce saw logs (Srivastava 1993), crop trees in the stand that meet criteria for pruning A. mangium timber production could supply a sub- (Beadle et al. 1994). Higher stockings also impose stantial part of the solid-wood market in the medium some control on average branch size. Large branches term. Knots and heart rot are undesirable characteris- are more difficult to prune and potentially more sus- tics that reduce the strength, appearance and value of ceptible to decay entry. A thinning strategy therefore the timber. Variable numbers of trees are also multi- becomes an inevitable part of the silviculture of stemmed at the base, the proportion probably related A. mangium plantations for solid-wood production. to genotype and site conditions (Srivastava 1993). Pruning 1 Ensis – the joint forces of CSIRO and Scion, Private Bag 12, Hobart, Tasmania 7001, Australia. Acacia mangium has persistent branches. This has Email: . led to the development of lift-pruning regimes where

79 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. branches are removed from the base of the tree Growth is less affected by pruning on a high than a upwards to convert the bottom log to clear or knot- low quality site (Pinkard and Beadle 1998a), so free wood (Mead and Speechly 1991; Weinland and pruning severity must be reduced on sites associated Zuhaidi 1991). The results of studies of a number of with slower rates of growth. In general, pruning species indicate that the preferred practice is green affects height growth less than it does diameter pruning that removes live rather than dead branches. growth. In a stand planted at 1100 stems per hectare Dead branches are associated with a high percentage in South Sumatra (C.L. Beadle, K. Barry, E. Hardi- of discoloration and decay in unpruned A. mangium yanto, R. Irianto, Junarto, C. Mohammed and A. (Ito and Nanis 1994). Pruning live branches also pre- Rimbawanto, unpublished data), height and diameter vents the development of loose knots and decreases growth following removal of 25% of crown length the size of the knotty core. using lift-pruning was significantly greater 18 The removal of live branches inevitably reduces months after pruning than in an unpruned control the productive capacity of an individual tree because (Table 2). However, the pruning treatment had been of the loss of green leaf area. This is of particular singled while the controls had not, and it is likely that concern in systems where only a proportion of the competition between multiple stems slowed indi- planted trees are selected for pruning, potentially vidual stem growth of the controls. putting them at a competitive disadvantage against In the same stand in South Sumatra, a form- the unpruned trees. Prescriptions must therefore opti- pruning treatment removed selected branches up to mise clear-wood production in such a way that there 3 m height that were either large or competing with is no significant effect on tree growth after pruning. the leader and until 25% leaf area had been removed Large branches are generally associated with poor from the tree. The effects on tree growth were not sig- stem form and with a high risk of decay entry after nificantly different from those observed with lift- pruning. In a study of fungal infection two years after pruning (Table 2). However, form-pruning was asso- pruning A. mangium, Weinland and Zuhaidi (unpub- ciated with improved stem straightness expressed as lished data; see Srivastava (1993)) noted that wounds reduced kink: more than 80% of the trees were of diameter >20 mm were always infected whether assessed as having slight or no kinks in the first 3 m the branches had been removed when green or dead. of the stem, the part that had been form-pruned In Eucalyptus nitens (Maiden), the length of decay (Table 3). Although current tree spacings at planting columns increased exponentially when the diameter and existing planting stock are inevitably associated of pruned branches was >20 mm. Branches at a with the development of some large branches (>30 narrow angle to the stem were also associated with a greater risk of decay (Mohammed et al. 2000). Thus, mm), pruning appeared to offer some level of control control of branch size is of vital importance in the sil- of their number in this experiment. viculture of plantations managed for solid wood. Table 1. The percentage of crown length removed Stocking and form-pruning provide two options for above which growth is reduced in a range of such control. Increasing the initial stocking density tree species pruned in plantations will reduce the incidence of large branches (Neilsen and Gerrand 1999) but more intense inter-tree com- Species % petition will then reduce the average growth of indi- Acacia mangiuma 40 vidual trees. Form-pruning selectively removes A. melanoxylonb 25 branches throughout the crown and can be used to reduce average branch size before subsequent lift- Eucalyptus grandisc 40 pruning (Pinkard 2002). E. nitensd 50 The proportion of branches that can be removed at Pinus patulae 25 any one pruning before growth is affected is a func- P. radiataf 35 tion of species and site (Table 1). Slower-growing P. sylvestrisg 40 species, for example, are more affected by pruning h than faster-growing species. In one experiment in Cryptomeria japonica 30 Malaysia, A. mangium tolerated the removal of 40% aMajid and Paudyal (1992); bMedhurst et al. (2003); cBredenkamp et al. (1980); dPinkard and Beadle (1998); of its crown length by lift-pruning before growth was eKarani (1978); fSutton and Crowe (1975); gLångström and significantly affected (Majid and Paudyal 1992). Hellqvist (1991); hFujimori and Waseda (1972)

80 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Table 2. Mean stem diameter, height increment and green crown lift of Acacia mangium 18 months after pruning at a plantation in South Sumatra. Means sharing the same letters are not significantly different at p<0.05. At age 18 months, when the trees were pruned (see text), their average height and diameter were 4.3 m and 4.1 cm, respectively, and there were no significant differences between treatments.

Pruning treatment Diameter increment (cm) Height increment (m) Green crown lift (m)

Control 6.83 a 7.44 a 2.49 a Form 8.28 b 8.35 b 3.07 a Lift 8.06 b 8.20 b 2.74 a

Green pruning triggers physiological responses or more lifts are required to ensure that crown that increase biomass production to a level that is sig- removal in any one lift does not exceed the level that nificantly greater than in a similar unpruned tree triggers a significant reduction in growth rate. While growing under the same environmental conditions. the compensatory responses just referred to have These physiological changes can be detected within a been demonstrated to work adequately for first-lift few days to weeks after pruning in eucalypts and aca- pruning in E. nitens and A. melanoxylon growing in a cias, and may be sustained for several months temperate climate (Pinkard and Beadle 2000; Med- (Pinkard and Beadle 2000; Medhurst et al. 2006). hurst et al. 2006), the physiological responses to sub- These changes in physiological activity are observed sequent lifts have not been as thoroughly as an increased rate of crown development (Pinkard investigated. The much higher rates of growth and Beadle 1998b) expressed through higher rates of observed in a tropical species mean that the elapsed leaf expansion, greater leaf development in the upper time between each lift-pruning will be a few months crown, greater leaf area to branch basal area ratio and only. In the experiment in South Sumatra referred to reduced leaf senescence. Significant increases in above, the base of the green crown was, on average, light-saturated rates of single-leaf net photosynthesis between 2.5 and 3.1 m above ground level 18 months are also observed to occur throughout the crown, after pruning (Table 2), and substantially beyond the though decrease in magnitude with depth in the height (≤1.5 m) to which branches had been pruned in canopy (Pinkard et al. 1998; Medhurst et al. 2006). the lift-pruning treatment.

Table 3. Percentage of assessed trees by kink class Thinning between 0–3 m height, 18 months after pruning at a plantation in South Sumatra. The Thinning serves to maximise the diameter growth of class codes for kink were: (1) no kinks, (2) trees by reducing intra-specific competition. In the slight kinks, (3) stem deviation stays within present context this is achieved by the harvesting or the centre line, (4) stem deviates outside the removal of unpruned trees. In A. mangium planta- centre line. The trees had been form-pruned tions managed for solid wood, the numbers of pruned to 3 m height at pruning (see text). trees in the final crop and the thinning strategy Pruning Percentage of trees by class code adopted will depend on the target tree-size and rota- treatment tion length. The commercial value of the thinned 1234 trees may also dictate the timing of the thinning oper- Control 25.0 36.2 19.4 19.4 ation. There are, however, some useful principles that Form 55.6 27.8 8.3 8.3 have emerged from thinning trials with other species Lift 25.0 36.1 13.9 25.0 that can help develop a workable strategy for thinning A. mangium. The implication of these findings is that pruning Thinning intensity affects stand growth. In a severity should be linked to the capacity of these E. nitens stand established at 1143 stems/ha (3.5 m × compensatory responses to result in no significant 2.5 m spacings), cumulative basal area growth per change in the growth of the pruned trees compared hectare was unaffected by thinning to 300 stems/ha with the unpruned trees in the stand. As pruning is seven years after thinning, while the removal of normally undertaken to a height of at least 4.5 m, two greater than 66% of standing basal area at thinning

81 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

(that resulted in a stand density of 100 stems/ha) was A pruning and thinning strategy associated with a significant reduction in cumulative basal area growth per hectare (Medhurst et al. 2001). The pruning and thinning strategy presented here Individual tree growth was improved with thinning uses current establishment practices for A. mangium. intensity and, in general, the dominants and co-dom- The pruning strategy is in part based on current prac- inants were the trees in the stand that produced a sig- tice used for A. mangium in Indonesia and Eucalyptus nificant basal area response to thinning. globulus and E. nitens in Australia (Gerrand et al. 1997), with the addition of form-pruning. As form- The growth response occurs because of changes in pruning to date has been used only as a research tool, the distribution of light and canopy photosynthesis in its benefits require further testing. The thinning the stand following thinning (Medhurst and Beadle strategy is linked to observations of current growth 2005). Significantly higher fractions of incident light rates of A. mangium in Australian and Indonesia. The are found in the middle and lower crowns of the trees timing of each operation attempts to meet the criteria and significantly greater light-saturated rates of pho- described above for optimising the quantity and tosynthesis are observed in the leaves of the lower quality of clear wood produced. crown, compared with unthinned stands. In the Acacia mangium plantations are established at results of a study by Medhurst and Beadle (2005), 3 × 3 m spacings (1111 stems/ha). The pruning and these changes were expressed through changes in the thinning strategy is then based on the following distribution of foliar nitrogen (N) content that result requirements: in positive relationships between N content and the • Form and lift-pruning to 4.5 m tree height, and fraction of incident light or light-saturated photosyn- thinning to reduce the stocking from 1111 to thetic rate. The increases in N content are related to a 296 stems/ha. significant decrease in specific leaf area (leaf area per • Four silvicultural interventions after planting, unit leaf weight; Medhurst and Beadle (2005)). excluding the need for any fertiliser application Changes in leaf area per tree are associated with an and weed control. increase in crown length in thinned stands (Medhurst • Every fifth row is an outrow removed at first 2000). thinning. These rows, equivalent to 222 stems/ha, These observations suggest that thinning at or soon are not used to select trees for pruning. after canopy closure will maximise the growth response: conversely, delaying thinning until after • The maximum number of trees available to select there has been appreciable crown lift will reduce the those for pruning is therefore 889 stems/ha (1111 – 222 = 889 stems/ha), equivalent to a magnitude of the response. Thinning intensity should selection ratio based on 296 stems/ha at final be commensurate with the maximisation of light stocking of about 1:3. interception, and therefore of growth, by individual trees. However, as thinning regimes aim to maximise Implementing the strategy is a four-step process stand production as well as optimise tree size, the (see Figure 1). ideal residual stand density after thinning will be that Step 1 which will lead to maximum leaf area index (leaf area Stage of growth: age 4–6 months per unit ground area) towards the end of the rotation, Operations or just ahead of the next thinning. This will depend, • Single trees of course, also on the tree sizes that are being sought Step 2 (Figure 1a) at harvest. Medhurst et al. (2001) found that, in general, the lower the quality of the site, the lower was Stage of growth: Mean total height @ 4 m the ability of the stand to respond to thinning. Thin- Operations ning intensity therefore needs to increase with • Form-prune two of every three trees of 889 trees, increasing site quality: smaller individual tree sizes i.e. 593 stems/ha, to 2.5 m (removes large branches are a given on lower-quality sites. As trees have a >15–20 mm diameter and high angle branches). capacity to develop longer tree crowns on high- The trees selected for form-pruning should be quality sites, delaying thinning will not, up to a point, those that meet criteria for good form (Table 4). preclude a growth response from thinning.

82 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Step 3 (Figure 1b,c) Step 4 (Fig. 1d) Stage of growth: Mean total height @ 8 m Stage of growth: Mean total height @ 12 m Operations Operations • Form-prune the best one of two trees pruned to • Lift-prune to 4.5 m trees that were form-pruned to 2.5 m (i.e. 296 of 593 stems/ha) to 4.5 m. 4.5 m. • Lift-prune these trees to 2.5 m. • Thin those trees that were form-pruned to 2.5 m • Thin all outrow trees (222 stems/ha) and the trees only (295 stems/ha). This will create the final that were not pruned at Step 2 (296 stems/ha). This stocking of 296 stems/ha. reduces the stocking from 1111 to 593 stems/ha).

(a) Step 2. (b) Step 3. Mean height @ 4 m Mean height @ 8 m

outrows

Figure 1. A representation of a pruning and thinning strategy for Acacia mangium. The trees are singled at age 4–6 months (see Step 1 in text). (a) In Step 2, two (black) out of every three trees are form-pruned to 2.5 m. (b) In Step 3, the better (blue) of each two trees form-pruned in Step 2 is now form-pruned to 4.5 m. The same trees are lift-pruned to 2.5 m. (c) Also in Step 3, the outrows (red) and the trees not pruned (also red) in Step 2 are thinned (harvested). (d) In Step 4, the trees form-pruned to 4.5 m are now lift-pruned to 4.5 m. The trees form-pruned only to 2.5 m in Step 2 are harvested. The final stocking rate is 296 stems/ha.

83 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

This strategy is based on form and lift-pruning to 10–12 years are assumed or inter-tree competition 4.5 m in height. Both are done in two sections (to will probably start to compromise individual tree- 2.5 m and then to 4.5 m), first the form-pruning then, growth rates. Thinning regimes that reduce final-crop after the tree has grown another 4 m in height, the lift- stocking to lower levels will then be required (Mead pruning is undertaken in the section that was previ- and Speechly 1991; Srivastava 1993) and will be ously form-pruned. The removal of branches >15–20 essential anyway if tree sizes >30 cm diameter breast mm diameter during form-pruning anticipates that height over bark are required. these branches would have been >30 mm diameter at lift-pruning. Care should be taken not to remove Heart rot more than 25–30% of the total leaf area during pruning at any of the above stages. Cut surfaces from pruning and singling are potential infection courts for the entry of decay-causing fungi Table 4. Criteria that together define acceptable form (Gales 2002; Lee 2002). Infection of heartwood by a for pruning an individual tree (adapted from range of white-rot fungi leads to heart rot, in which Beadle et al. (1994)). These are used to the decayed wood appears fibrous and stringy as well ensure that the better trees are selected for pruning. If insufficient trees required for as a pale yellowish-white (Lee et al. 1988). Whereas clear wood meet these criteria, it may be heart rot can be tolerated in wood destined for pulp, it necessary to select some trees that do not is not acceptable in solid-wood products. meet all the criteria. Form-pruning is a Heart rot has been recorded in pruned plantations mechanism for increasing the number of of A. mangium during a number of studies in trees that will eventually meet the criteria. Malaysia (Lee et al. 1988; Ito 2002). Unhealed pruning wounds have been recorded as contributing a 1 Single-stemmed and free of secondary leaders to up to 62.5% of heart rot infections (Lee et al. 2 Straight stem with minimal stem deformations from the vertical 1988). During a survey of heart rot in A. mangium 3 Stems free from wounds and disease plantations in Indonesia, Barry et al. (2004) found the 4 No branches already >30 mm in diameter highest incidence of heart rot was associated with a 5 Butt sweep limited to the bottom 0.3 m region where pruning occurred. In contrast, Zakaria a Secondary leaders that form above the final pruning height et al. (1994) found no relationship between the inci- can be acceptable dence of heart rot and pruning of A. mangium in Malaysia. In a study in South Sumatra, no heart rot Two thinning operations are also required, one was detected 18 months after pruning (Beadle et al., when the average tree height is about 8 m, the second unpublished data). In an A. mangium provenance when tree height is about 12 m. The size of the trial, also conducted in South Sumatra, Suberanjeriji- thinned trees may be suitable for pulpwood but care plus (seed stock similar to that used by Beadle et al.) should be taken not to damage retained trees during trees were found to have the second lowest incidence the thinning operations. This is assisted by the crea- of heart-rot infection associated with drill wounds of tion of an outrow at the first thinning to help access to a total of six provenances (Barry et al. 2006). Thus, the other rows for individual tree removal. In one while pruning has been linked to increased heart rot sense the strategy is conservative in that it may not be in some regions in Indonesia (Barry et al. 2004) and necessary in Step 2 to form-prune twice the number Malaysia (Lee et al. 1988) it appears to be of impor- of trees that are going to be retained in the final crop: tance only when the plant material is susceptible and the additional form-pruning is a cost but it allows a sufficient source of heart rot fungi is present to greater choice of the final-crop trees at Step 3. How- invade the wounds. ever, the timing of the thinning operation is to some extent speculative and assumes that each of the two Synthesis operations will occur before canopy lift commences in the pruned trees. The required size of the final-crop While the fundamental requirements of green trees will determine the time of harvest. Given the pruning and thinning in plantations managed for types of growth rates experienced in A. mangium solid wood are well established, the development of plantations, rotation lengths of no longer than about pruning and thinning schedules for Acacia mangium

84 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. remain in its infancy. Form-pruning appears to offer Development and Training Project, study report. Sabah, an approach for improving stem straightness. How- Chin Chi Printing Works, 52p. ever, its coordination with subsequent lift-pruning Karani, P.K. 1978. Pruning and thinning in a Pinus patula requires further investigation. How to schedule the stand at Lendu plantation, Uganda. Commonwealth timing of thinning operations is a complex issue. The Forestry Review, 57, 269–278. first requirement is that their timing should ensure Långström, B. and Hellqvist, C. 1991. Effects of different that branches remain green but at the same time do pruning regimes on growth and sapwood area of Scots not become large. The second requirement is that the pine. Forest Ecology and Management, 44, 239–254. timing of thinning should aim maximise tree growth Lee, S.S., 2002. Overview of the heartrot problem in Acacia rates, stand production and the value of the thinned as — gap analysis and research options. In: Barry, K., ed., well as the final-crop trees. Just how to manage what Heartrots in plantation hardwoods in Indonesia and Australia. Canberra, ACIAR Technical Reports, No. 51e, can be difficult compromises should also be a focus 26–34. for new research. Lee, S.S., Teng, S.Y., Lim, M.T. and Kader, R.A. 1988. Discoloration and heartrot of Acacia mangium Willd. — References some preliminary results. Journal of Tropical Forest Science, 1, 170–177. Barry, K.M., Irianto, R.S.B., Tjahjono, B., Tarigan, M., Majid, N.K. and Paudyal, B.K. 1992. Pruning trial for Agustini, L., Hardiyanto, E.B. and Mohammed, C.L. Acacia mangium Willd. plantation in Peninsular 2006. Variation of heartrot, sapwood infection and Malaysia. Forest Ecology and Management, 47, 285– polyphenol extractives with provenance of Acacia 293. mangium. Forest Pathology, 36, 183–197. Mead, D.J. and Speechly, H.T. 1991. Growing Acacia Barry, K.M., Irianto, R.S.B., Santoso, E., Turjaman, M., mangium for high quality sawlogs in Peninsular Widyati, E., Sitepu, I. and Mohammed, C.L. 2004. Malaysia. In: Sheikh Ali Abod et al., ed., Recent Incidence of heart rot in harvest-age Acacia mangium in developments in tree plantations of humid/sub-humid Indonesia using a rapid survey method. Forest Ecology tropics of Asia. Serdang, Selangor, Universiti Pertanian and Management, 190, 273–280. Malaysia, 54–69. Beadle, C.L., Turnbull, C.R.A. and McLeod, R. 1994. An Medhurst, J.L. 2000. Growth and physiology of Eucalyptus assessment of growth and form for pruning to 6 metres in nitens in plantations following thinning. PhD thesis, E. nitens plantations. Tasforests, 6, 1–6. University of Tasmania, Australia. Bredenkamp, B.V., Malan, F.S. and Conradie, W.E. 1980. Medhurst, J.L. and Beadle, C.L. 2005. Photosynthetic Some effects of pruning on growth and timber quality of capacity and foliar nitrogen distribution in Eucalyptus Eucalyptus grandis in Zululand. South African Forestry nitens is altered by high-intensity thinning. Tree Journal, 114, 29–34. Physiology, 25, 981–991. Fujimori, T. and Waseda, O. 1972. Fundamental studies on Medhurst, J.L., Beadle, C.L. and Neilsen, W.A. 2001. pruning II. Effects of pruning on stem growth. Bulletin of Growth in response to early-age and later-age thinning in the Government Experimental Station, 244, 1–15. Eucalyptus nitens (Deane and Maiden) Maiden Gales, A., 2002. Heartrot in plantations — significance to plantations. Canadian Journal of Forest Research, 31, the wood processing industry. In: Barry, K., ed., Heartrots 187–197. in plantation hardwoods in Indonesia and Australia. Medhurst, J.L., Pinkard, E.A., Beadle, C.L. and Worledge, Canberra, ACIAR Technical Reports No. 51e, 18–21. D. 2003. Growth and stem form responses of plantation- Gerrand, A.M., Neilsen, W.A. and Medhurst, J.L. 1997. grown Acacia melanoxylon (R. Br.) to form-pruning and Thinning and pruning eucalypt plantations for sawlog nurse-crop thinning. Forest Ecology and Management, production in Tasmania. Tasforests, 9, 15–34. 179, 183–193. Hardiyanto, E. 2006. Options for solid-wood products from Medhurst, J.L., Pinkard, E.A., Beadle, C.L. and Worledge, Acacia. In: Potter, K., Rimbawanto, A. and Beadle, C., D. 2006. Increases in photosynthetic capacity of ed., Heart rot and root rot in tropical Acacia plantations. plantation-grown Acacia melanoxylon after form- Canberra, ACIAR Proceedings No. 124, 51–56. [These pruning. Forest Ecology and Management, in press. proceedings] Mohammed, C., Barry, K., Battaglia, M., Beadle, C., Eyles, Ito, S., 2002. The incidence of heartrot and disease severity A., Mollon, A. and Pinkard, E. 2000. Pruning-associated on several Acacia species in SAFODA plantations. Study stem defects in plantation E. nitens and E. globulus grown report of SAFODA–JICA project, 16p. for sawlog and veneer in Tasmania. In: The future of Ito, S. and Nanis, L.H. 1994. Heartrot of Acacia mangium in eucalypts for wood production. Proceedings of IUFRO SAFODA plantations. Sabah Reafforestation Technical Conference, Launceston, Tasmania, Australia, 357–364.

85 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Neilsen, W.A. and Gerrand, A.M. 1999. Growth and Rimbawanto, A., 2002. Plantation and tree improvement branching habit of Eucalyptus nitens at different spacing trends in Indonesia. In: Barry, K., ed., Heartrots in and the effect on final crop selection. Forest Ecology and plantation hardwoods in Indonesia and Australia. Management, 123, 217–229. Canberra, ACIAR Technical Reports, No. 51e, 18–21. Pinkard, E.A., 2002. Effects of pattern and severity of Srivastava, P.B.L. 1993. Silvicultural practices. In: Awang, pruning on growth and branch development of pre- K. and Taylor, D. ed., Acacia mangium, growing and canopy closure Eucalyptus nitens. Forest Ecology and utilization. FAO, Bangkok, Thailand, and Winrock Management, 157, 217–230. International, MPTS Monograph Series No. 3, 113–147. Pinkard, E.A. and Beadle, C.L. 1998a. Effects of green Sutton, W.R.J. and Crowe, J.B. 1975. Selective pruning of pruning on growth and stem shape of Eucalyptus nitens. radiata pine. New Zealand Journal of Forest Science, 5, New Forests, 15, 107–126. 171–195. — 1998b. Above-ground biomass partitioning and crown Weinland, G. and Zuhaidi, A.Y., 1991. Management of architecture of Eucalyptus nitens (Deane and Maiden) Acacia mangium stands: tending issues. In: Appanah, S., Maiden following green pruning. Canadian Journal of Forest Research, 28, 1419–1428 Ng, F.S.P. and Ismail, R., ed., Malaysian forestry and forest products research proceedings. Kuala Lumpur, Pinkard, E.A. and Beadle, C.L. 2000. A physiological Forest Research Institute Malaysia, 40–52. approach to silvicultural pruning. International Forestry Review, 2, 295–305. Zakaria, I., Wan Razali, W.H., Hashim, M.N. and Lee, S.S. Pinkard, E.A., Beadle, C.L. Davidson, N.J. and Battaglia, 1994. The incidence of heart rot in Acacia mangium M. 1998. Photosynthetic responses of Eucalyptus nitens plantations in Peninsula Malaysia. Forest Research (Deane and Maiden) Maiden to green pruning. Trees: Institute Malaysia, Research Pamphlet No. 114, 15p. Structure and Function, 12, 119–129.

86 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

Options for solid-wood products from Acacia mangium plantations

Eko B. Hardiyanto1,2

Abstract

Acacia mangium wood has proved to be not only suitable for producing high-quality pulp and paper, but also an excellent material for solid-wood products. There has been growing interest in utilising wood of A. mangium for solid wood, corresponding with the declining availability of logs from native forests. The currently available sawlogs of A. mangium are harvested from unpruned and unthinned plantations, and consequently are of low quality (many knots and poor stem form) with poor recovery of sawn timber. Proper silvicultural techniques that incorporate pruning and thinning are of crucial importance in growing plantations for solid wood. Acacia mangium can be grown for solid-wood products with a rotation of around 10 years that is expected to produce a total stem volume more than 200 m3 per ha: about 30% of it will be for solid wood. Minimum tree diameter at breast height will be 30 cm. The current price of sawlogs of A. mangium (from unpruned and unthinned plantations) is considered low, but it is expected to increase with reducing availability of logs from natural forest, increased familiarity of its users with the wood, and better log quality from plantations managed specifically for solid-wood products.

Since the early 1990s, industrial forest plantations pulp and paper, but also good for wood products such have been developed in Indonesia. As of 2004, a total as plywood, furniture, flooring and light construc- of 2,500,966 ha of pulp plantation and 865,256 ha of tion. Lately there has been growing interest in uti- sawlog plantation had been established (Dirjen BPK lising wood of A. mangium for solid-wood products. 2005). Acacia plantations have been established pri- This corresponds with the reduced availability of marily to produce wood for the pulp and paper indus- logs from native forests. The continuing pressure to tries. Acacia mangium Willd. is the main plantation protect native forest will further enhance the oppor- species planted on mineral soils, while Acacia crassi- tunity to develop sawlog plantations of A. mangium. carpa A. Cunn. ex Benth. is the only species grown on peat land. The pulp properties of A . mangium Solid-wood utilisation wood are comparable to those of many Eucalyptus species. A number of studies on the utilisation of A. Various studies have been conducted to examine the mangium show that its wood is not only excellent for suitability of A. mangium for products other than pulp and paper. The density of A. mangium wood varies from 420 to 483 kg/m3. It is considered to be 1 PT. Musi Hutan Persada, PTC Mall Blok I-9, Jl. R. stable, with shrinkage from fresh to air dry of around Sukamto, Palembang 30114, Indonesia. 2 Faculty of Forestry, Gadjah Mada University, 6.4% tangentially and 2.7% radially (Abdul-Kader Bulaksumur, Yogyakarta 55281, Indonesia. and Sahri 1993). Its fibre is relatively straight and Email: ; . grain (Yamamoto 1998).

87 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124.

The timber of A. mangium dries well, fairly rapidly bawanto, unpublished data). In areas where the inci- and without serious defects when a suitable kiln dence of heart rot is low, the development of sawlog schedule is used (Abdul-Kader and Sahri 1993). plantations is considered to be possible. However, the wood is prone to initial checking and Knots are the major factor limiting the utilisation can also collapse and shrink during drying, resulting of solid wood harvested from unpruned stands of A. in substantial losses (Yamamoto 1998), hence the mangium. Consequently, unless silvicultural tech- need for a proper kiln schedule to obtain good-quality niques are used to control branching and hence the timber. occurrence of knots, the quality of wood from this The wood of A. mangium is classified as light hard- species may limit its utilisation. The recovery rate of wood with low to moderate strength. It has good sawn timber board from sawlogs harvested from machining properties and is suitable for making furni- unpruned and unthinned A. mangium stands is low; ture, cabinets, mouldings, and door and window com- about 38% at PT. Musi Hutan Persada (Supriyadi, ponents (Abdul-Kader and Sahri 1993). The wood is pers. comm. 2005) and 35% in S. Kalimantan (Thorp also suitable for light structural works, agricultural 2005), for example. By applying specific silvicul- implements, boxes and crates (Abdul-Kader and Sahri tural treatments, there is the potential to substantially 1993; Yamamoto 1998). A recent study on the increase the recovery rate of A. mangium sawlogs. mechanical properties of sawn timber indicated that Another defect that may reduce the general quality the wood of A. mangium can be used for construction of the end product and limit the usefulness of the and meets the requirements of the Indonesian Standard A. mangium sawlogs is end-splitting, particularly of for Wood Construction (Amalia 2003; Firmanti and trees harvested from stands under 10 years old. End- Kawai 2005). The wood of A. mangium can also be splitting is a phenomenon manifested during felling easily processed into veneer and plywood. The peeling and cross-cutting of logs; it is caused by high growth process is considered easy and the green veneers pro- stress which normally occurs in fast-growing tree spe- duced are tight, smooth and of acceptable quality cies, particularly hardwoods. Growth stress is gener- (Abdul-Kader and Sahri 1993; Yamamoto 1998). ated within woody tissue as a result of the tendency of Heart-rot disease in A. mangium has been reported differentiating cells to contract during cell maturation in a number of studies and can reduce the utilisation in the longitudinal direction and expand in the trans- of solid wood since heart rots reduce timber volume verse direction (Malan 1995). In Eucalyptus grandis, and quality (Lee et al. 1988; Lee 2002; Ito 2002). growth stress is reported to be highly heritable, and A recent survey conducted at a number of sites in has no relationship with other characteristics such as Indonesia revealed that the incidence of heart rot diameter, tree height and wood density (Malan 1995). varied according to site, from low (6.7% East Kali- Observations of logs harvested from A. mangium mantan, 11.3% South Sumatra) to reasonably high trees of more than 10 years of age revealed that the (35.3% Jambi, 46.7% West Java). A combination of proportion of logs showing end-splitting was small. differing plantation management strategies—for example, pruning, age and local conditions— Silviculture of plantation for explained the differences in heart-rot incidence (Barry et al. 2004). Pruning was reported to be linked solid-wood products to increased incidence (Barry et al. 2004), but may be Establishment of importance only when the plant material is susceptible and a sufficient source of heart-rot fungi is The method of plantation establishment of A. present in the environment to invade the wounds mangium developed for solid-wood products is caused by pruning. In addition, while wounds caused similar to that for plantations grown for pulp. The site by singling are typically assumed to have a large is generally prepared manually by blanket spraying impact on the incidence of heart rot, an influence of existing unwanted vegetation with herbicide. Seed- singling on heart-rot incidence in a provenance trial lings raised in the nursery are planted manually at an in the Riau province of Sumatra was not apparent initial stocking rate of 1100/ha. Genetically improved (Barry et al. 2006). A pruning study in South Sumatra seeds should be used for growing sawlog plantations. also detected no heart rot 18 months after singling Trees should be fertilised using a basal fertiliser of and pruning (C. Beadle, K. Barry, E.B. Hardiyanto, phosphorus at planting time, usually in the planting R. Irianto, Junarto, C. Mohammed and A. Rim- hole. Nitrogen fertiliser may not be required in the

88 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. second rotation as a lack of a positive response is the first thinning when trees are around 8–9 m tall. The likely due to high nitrogen content available in the soil last pruning should be carried out at 4–5 years of age to as a result of the fixing of atmospheric nitrogen by the a pruning height of 6 m. The minimum length of previous plantation (Hardiyanto et al. 2004). sawlog is 4 m (Hardiyanto 2004). Reducing weed competition is of crucial importance As mentioned previously, A. mangium has poor in the early phase of plantation establishment so that apical dominance and tends to have large branches. In trees can grow optimally and close their canopy in the a study of fungal infection 2 years after pruning, first year. Weeds are normally controlled by hand- Weinland and Zuhaidi (1992) noted that wounds of weeding at age 3–4 months followed by a second diameter >20 mm were always infected whether the hand-weeding and herbicide application at about 6 branches had been removed when green or dead. months of age. Depending on the weed growth, the Therefore, control of branch size using form-pruning hand-weeding may be reapplied at age 12 months. is vital in the silviculture of plantations managed for solid wood. Unlike lift-pruning, form-pruning selec- Singling and pruning tively removes branches throughout the crown and can be used to reduce average branch size before a In the establishment of plantations for solid wood subsequent lift-pruning (Pinkard 2002) or to correct it is of paramount importance to have early silvicul- potential deviation of stems from a pathway of ver- tural intervention that utilises pruning and thinning. tical growth (Medhurst et al. 2003). A pruning trial The timing and method of pruning and thinning is conducted at an 18-month-old stand of A. mangium at crucial. As A. mangium has weak apical dominance PT. Musi Hutan Persada’s plantation found that form- and is inherently multi-stemmed, singling is neces- pruning reduced the number of trees with large sary. Singling to cull and leave one of the best stems branches and increased the number of trees with better is carried out when trees are around 3–4 months old. stem form (reduced the number of stem kinks) Unlike plantations grown for pulp, plantations to be (C. Beadle et al., unpublished data). In the same trial, utilised for solid-wood have to be pruned to produce lift and form-pruning were found to have no associa- high-quality logs. This is important, as A. mangium tion with heart rot (C. Beadle et al., unpublished data). does not shed dead lower branches naturally and often the dead branches persist for a long time and are associated with deterioration in wood quality (Malan 1995; Thinning Waugh 1996). Green and dead knots caused by branching reduce sawn timber yields. Dead branches Thinning involves the removal of part of the stand result in loose knots that fall out to leave holes in the in order to concentrate future volume growth on sawn timber. Dead branches are also associated with a fewer and better-quality stems. The advantage of high percentage of discoloration and decay (Ito and thinning is that trees left behind have greater Nanis 1994). The objective of pruning is therefore to resources (water, light, space to grow) and therefore maximise the amount of clear wood produced by a increase in size more rapidly. Thus, by selecting the tree. If branches are properly pruned while green, then best trees for retention and enabling those to grow there is a high probability that new wood will grow faster, the rate of increase of stand value increases. over the pruned branch stubs and that knot-free clear Thinning improves stand quality through the wood will be grown on the stem. Green pruning, that removal of deformed trees and a reduction in the time is, removal of leaf area, has the potential to decrease taken for trees to reach valuable sawlog size. Thin- tree growth rates, particularly stem diameter. A study ning is the most powerful tool to manipulate the on pruning of A. mangium revealed that removing the development of the plantation and the quality of the crown below 50% of tree height depressed growth sig- final stand. Thinning increases timber revenue by nificantly, especially stem diameter (Majid and increasing the volume of sawlog produced. This is Paudyal 1992). The first lift-prune, to 40% of the tree because larger trees attract significantly higher height, should therefore be carried out when trees are prices, as they are less expensive to harvest (on a Rp/ about 6 months old and about 2.5–3.0 m tall. This m3 basis) and yield more-valuable end products. should avoid a significant reduction in stem diameter Acacia mangium is very competitive for light, growth. The second pruning, to a height of 4 m, should moisture and nutrients and it is important that it main- be conducted at 1.5–2.0 years old, at the same time as tains active growth. Early thinning allows the best

89 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. trees in the stand to take advantage of any improved steadily in recent years as the industry gains more growing conditions. To ensure this, the first thinning confidence in its availability and experience in how should be conducted when the majority of trees are to use it, and wood supplies from natural forest con- around 8–9 m in height; this should occur at 1.5–2.0 tinue to decline. years of age, with overall stocking reduced to 500–550 The increasing demand for wood from plantations, trees/ha. The remaining trees can then maintain good including that of A. mangium, will likely cause a rise growth until about age 4–5 years when inter-tree com- in log prices and the investment in plantations for petition resumes. The second thinning is then carried solid-wood utilisation will become economically out to leave 200–250 trees/ha. The remaining trees are viable. Current prices of harvested sawlog from grown on until the final harvest at around age 10 years. unpruned and unthinned A. mangium stands are con- The growth data reported from A. mangium stands pre- sidered low due to low log quality (many knots and viously thinned show that at about 10 years of age the poor stem form). Logs harvested from managed average diameter at breast height is more than 30 cm sawlog plantations are expected to have better sawlog and the average height was more than 26 m. About yield, wood quality and product value and command 30% of the total stem volume can be utilised for higher prices. Nevertheless, the economic aspects of sawlog while the remaining wood can be used for pulp growing plantations for solid wood (growing cost and (Figure 1) (Hardiyanto and Supriyadi 2005). value of logs) need to be assessed and compared with those of growing pulp plantations Economic prospects Future research and development At present it is difficult to estimate the future demand for A. mangium timber, as its milling, kiln drying and As mentioned in the preceding section, the develop- related characteristics are still being assessed. How- ment of A. mangium plantations for solid wood is still ever, demand for A. mangium timber has increased in its infancy and therefore much research work has

Figure 1. A 10-year-old, thinned stand of Acacia mangium in Merbau, South Sumatra having average stem diameter, height and total stem volume of 36.6 cm, 26 m and 228.7 m3/ha, respectively.

90 From: Potter, K., Rimbawanto, A. and Beadle, C., ed., 2006. Heart rot and root rot in tropical Acacia plantations. Proceedings of a workshop held in Yogyakarta, Indonesia, 7–9 February 2006. Canberra, ACIAR Proceedings No. 124. yet to be done. The following areas need attention in Hardiyanto, E.B. 2004. Silvikultur dan pemuliaan Acacia the future research and development of A. mangium mangium. Dalam: Hardiyanto dan Arisman, H., ed., plantations for solid-wood products: Pembangunan hutan tanaman Acacia mangium- • genetic improvement — to improve growth, pengalaman di PT. Musi Hutan Persada, Sumatera Selatan. PT. 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